Ojima I. (ed.) Fluorine in Medicinal Chemistry and Chemical Biology

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318 Fluorine in Medicinal Chemistry and Chemical Biology

the basicity of the amino group due to the strong electron - withdrawing nature of

ﬂ uorine.

Taguchi and co - workers synthesized several biologically active gem -

diﬂ uorocyclopropanes, such as metabotropic glutamate receptor agonist 12 [15] and

methylene - gem - diﬂ uorocyclopropane 13 [16] . 2 - (2 - Carbohydroxy - 3,3 - diﬂ uorocyclopropyl) -

glycines (F

CCGs) were synthesized by means of a chiral auxiliary method (for F

CCG - I

12 [15a, 15c] ) as well as a chiral pool method (for all eight possible stereoisomers [15c] ).

Enhancement of acidity of the ω - carboxyl group by ﬂ uorine incorporation was conﬁ rmed

by NMR titration experiments. Evaluation of the neuropharmacological activities of

CCGs in rat, compared with those of the corresponding cyclopropylglycine isomers,

revealed that the (2 S ,1 ′ S ,2 ′ S ) - isomer, l - F

CCG - I 12 , was a potent agonist for metabotropic

Figure 12.6 Examples of biologically active gem - diﬂ uorocyclopropanes (simple

molecules).

gem-Diﬂ uorocyclopropanes as Key Building Blocks for Novel Biologically Active Molecules 319

glutamate receptors, which induced a priming effect on α - aminopimelate and l - glutamate

[15c, 15d, 5e] . Methylenecyclopropylglycine (MCPG) and its metabolite, MCPF - CoA,

are known to have inhibitory activity against enoyl - CoA hydratase (crotonase), respon-

sible for fatty acid metabolism [17] . Four possible stereoisomers of methylene - gem -

diﬂ uorocyclopropylglycines 13 (F

MCPG) were synthesized [16] , although their

biological activities were not reported.

Barger and co - workers also synthesized F

MCP derivatives 14 and evaluated their

activities against the nematode pest Meloidogyne incognite [18] . Unfortunately, com-

pounds 14 were inactive up to a concentration of 400 ppm, while the corresponding non-

ﬂ uorinated compound (R = H) showed 100% inhibition of the microbe at only 7.8 ppm

[18] . Kirihara et al. reported a highly efﬁ cient method for the synthesis of the optically

active gem - diﬂ uorocyclopropane analogue 15 of 1 - aminocyclopropane - 1 - carboxylic acid

using a chemo - enzymatic strategy [19] . Kirk, Haufe, and their co - workers synthesized

2,2 - diﬂ uoro - 1 - phenylcyclopropylamine ( 16 ) and evaluated its inhibitory activity for

ﬂ avin - containing monoamine oxidase (MAO). Unfortunately, no inhibitory effect was

observed and it was concluded that monoﬂ uorine substitution was essential to cause MAO

inhibition [20] . Recently, Roe et al. reported an interesting biologically active simple gem -

diﬂ uorocyclopropane 17 , which acted as a competitive inhibitor of the juvenile hormone -

epoxide hydrolase [21] .

We developed a synthetic methodology for preparing optically active gem -

diﬂ uorocyclopropane building blocks using lipase technology [4] . Using this methodology,

gem - diﬂ uorocyclopropanes 18 and 19 , bearing a 9 - anthracenecarbonyl group, were syn-

thesized. These compounds showed a strong DNA - cleavage property switched on by

photoirradiation [22] (see Figure 12.6 ).

Nucleoside analogues have been recognized as potential chemotherapeutic agents and

in fact several carbocyclic nucleosides exhibit potent anti - HIV activities. Accordingly,

gem - diﬂ uorocyclopropane - containing nucleoside analogues and mimics have attracted

substantial interest and various compounds have been synthesized (see Figures 12.7

and 12.8 ). Csuk pioneered this ﬁ eld and his group prepared a good number of gem -

diﬂ uorocyclopropane - containing nucleoside mimics as represented by compounds 20 – 25

[23] , some which were found to possess antitumor activity and anti - HIV activity. Although

all compounds reported by Csuk ’ s group are racemic, optically active compounds could

be prepared using the enzymatic resolution technology developed by us [4] and by

Kirihara ’ s group [19] .

Zemlicka and co - workers synthesized methylene - gem - diﬂ uorocyclopropane mimics

of nucleoside and found that the Z - isomer of adenine - 9 - yl analogue 26 exhibited anticancer

activity against leukemia and solid tumor cell lines in vitro (see Figure 12.8 ) [24] .

Adenin - 9 - yl, 2 - amino - 6 - chloropurin - 9 - yl, and guanin - 9 - yl analogues of 26 were synthe-

sized and evaluated for their activities against HCMV, HSV - 1, HSV - 2, EBV, VZV, and

HIV - 1. However, only the E - isomer of adeninyl analogue 26 showed moderate antiviral

activity against the Towne strain of HCMV propagated in human foreskin ﬁ broblast (HFF)

cells (EC

21 µ M), which was noncyctotoxic at this concentration [24] . Recently, Robins

and co - workers reported the synthesis of the gem - diﬂ uorocyclopropane analogues of

nucleosides such as 27 and 28 (see Figure 12.8 ) [25] , but no biological activities of these

compounds have yet been disclosed.

320 Fluorine in Medicinal Chemistry and Chemical Biology

Figure 12.8 Nucleoside - type biologically active gem - diﬂ uorocyclopropanes prepared by the

Zemlick and Robins group.

Figure 12.7 Nucleoside - type biologically active gem - diﬂ uorocyclopropanes prepared by the

Csuk group.

gem-Diﬂ uorocyclopropanes as Key Building Blocks for Novel Biologically Active Molecules 321

12.3 Synthesis of gem - Diﬂ uorocyclopropanes

Three synthetic methods for gem - diﬂ uorcyclopropane derivatives have been developed

to date. Shibuya and Taguchi reported an efﬁ cient route to optically active gem -

diﬂ uorocyclopropane derivative 30a through diastereoselective Michael addition of Evans

enolate 29a to 4 - bromo - 4,4 - diﬂ uorobut - 2 - enoate, followed by Et

B - mediated radical cou-

pling (see equation 1, Scheme 12.1 ) [15a] . Alternatively, for the synthesis of metabotropic

glutamate receptor agonist F

CCG - I, the Michael acceptor having the Evans oxazolidinone

as the chiral auxiliary was reacted with N - diphenylmethyleneglycinate 29b in DMF to

give diﬂ uorocyclopropylglycine derivative 30b with an excellent diastereoselectivity

( > 95% de) (see equation 2, Scheme 12.1 ) [15c] .

The most versatile method for constructing gem - diﬂ uorocyclopropanes is the

carbine - addition reaction to oleﬁ ns, which includes two types as shown in Scheme 12.2

(type A and type B). Boger and Jenkins synthesized the gem - diﬂ uorocyclopropane moiety

through intramolecular metal - carbenoid addition to 1,1 - diﬂ uoroalkene group (type A).

Thus, diazoquinone 31 was treated with 0.1 – 0.2 equivalents of Rh

(OAc)

in toluene at

110 ° C to give gem - diﬂ uorocyclopropane 32 in good yield [8] (see equation 3 in Scheme

12.2 ). The most widely used method for preparing gem - diﬂ uorocyclopropanes is the

diﬂ uorocarbene addition to oleﬁ ns (type B), which is applicable to large - scale preparation.

Although numerous reactions for generating diﬂ uorocarbene have been reported, three

major methods are shown under type B in Scheme 12.2 [26 – 29] : (a) pyrolysis of chloro-

diﬂ uoroacetate [26] or trimethylsilyl ﬂ uorosulfonyldiﬂ uoroacetate (TFDA), known as

Dolbier ’ s reagent [27] ; (b) generation from dibromodiﬂ uoromethane following Barton ’ s

Scheme 12.1 Synthesis of gem - diﬂ uorocyclopropane through diastereoselective Michael

reaction followed by radical coupling.

322 Fluorine in Medicinal Chemistry and Chemical Biology

or Schlosser ’ s procedure [28] ; and (c) generation from triﬂ uoromethylmercury compounds,

known as Seyferth ’ s reagents [29] . Of these methods, (b) and (c) can generate diﬂ uoro-

carbenes under mild conditions. Unfortunately, CF

and Seyferth ’ s reagent are no

longer commercially available because of their inherently hazardous properties. Therefore,

ClCF

COONa is currently the sole commercial source for diﬂ uorocarbene generation.

Since this reaction requires harsh conditions (180 – 190 ° C in diglyme), development of a

new and efﬁ cient method is desirable for the generation of diﬂ uorocarbene under mild

conditions.

Scheme 12.3 summarizes typical examples for the synthesis of gem -

diﬂ uorocyclopropanes via diﬂ uorocarbene addition to oleﬁ ns. Although high temperature

is necessary to generate diﬂ uorocarbene by pyrolysis of ClCF

COONa, addition of

diﬂ uorocarbene to carbon – carbon double bonds proceeds in a stereospeciﬁ c cis manner

Scheme 12.2 Preparation of gem - diﬂ uorocyclopropanes through addition of diﬂ uorocarbene

to oleﬁ n.

gem-Diﬂ uorocyclopropanes as Key Building Blocks for Novel Biologically Active Molecules 323

Scheme 12.3 Typical examples of the synthesis of gem - diﬂ uorocyclopropanes using diﬂ uo-

rocarbene addition to oleﬁ n.

324 Fluorine in Medicinal Chemistry and Chemical Biology

[19a] . Taguchi and co - workers prepared optically pure gem - diﬂ uorocyclopropane 34

through addition of diﬂ uorocarbene to optically active oleﬁ n ( S ) - 33 , followed by

chromatographic separation of the resulting diastereomers, ( S,R,R ) - 34 and ( S,S,S ) - 34

(2.6 : 1, 77% yield) (see equation 4, Scheme 12.3 ) [15a] . We prepared novel bis - gem -

diﬂ uorocyclopropanes 36 as a mixture of dl - 36 and meso - 36 (1 : 1.5, 81% yield) through

diﬂ uorocarbene addition to ( E,E ) - diene 35 [4b] (see equation 5, Scheme 12.3 ). Generally,

pyrolysis of trimethylsilyl ﬂ uorosulfonyldiﬂ uoroacetate (TFDA) proceeds under milder

conditions (130 ° C) as shown in equation 6, Scheme 12.3 [14] . However, TFDA is avail-

able only in a few countries at present. Diﬂ uorocarbene can be generated at room tem-

perature under the Barton – Schlosser conditions (CF

, PPh

, KF, and 18 - crown - 6).

Interesting spiro - gem - diﬂ uorocyclopropane 40 was synthesized by de Meijere and co -

workers using this method (see equation 7, Scheme 12.3 ) [30] . To the best of our knowl-

edge, this is the most efﬁ cient method for preparation of gem - diﬂ uorocyclopropanes.

However, as mentioned above, it is impossible to obtain hazardous CF

commercially.

For experienced researchers using appropriate safety precautions, toxic Seyferth ’ s reagent

is a useful source for preparing gem - diﬂ uorocyclopropanes. Robins and co - workers syn-

thesized gem - diﬂ uorocyclopropane nucleoside 42 using this method because substrate 41

was not tolerant of high temperature (see equation 8, Scheme 12.3 ) [25] .

12.4 Synthesis of Optically Active gem - D i ﬂ uorocyclopropanes via

Enzymatic Resolution

Enzymatic resolution has been successfully applied to the preparation of optically active

gem - diﬂ uorocyclopropanes (see Scheme 12.4 ). We succeeded in the ﬁ rst optical resolution

of racemic gem - diﬂ uorocyclopropane diacetate, trans - 43 , through lipase - catalyzed

enantiomer - speciﬁ c hydrolysis to give ( R,R ) - ( − ) - 44 with > 99% ee (see equation 9, Scheme

12.4 ) [4a] . We also applied lipase - catalyzed optical resolution to an efﬁ cient preparation

of monoacetate cis - 46 from prochiral diacetate cis - 45 (see equation 10, Scheme 12.4 ) [4a] .

Kirihara et al. reported the successful desymmetrization of diacetate 47 by lipase - catalyzed

enantiomer - selective hydrolysis to afford monoacetate ( R ) - 48 , which was further trans-

formed to enantiopure amino acid 15 (see equation 11, Scheme 12.4 ) [19] . We demon-

strated that the lipase - catalyzed enantiomer - speciﬁ c hydrolysis was useful for

bis - gem - diﬂ uorocyclopropane 49 . Thus, optically pure diacetate ( R,S,S,R ) - 49 and ( S,R,R,S ) -

diol 50 , were obtained in good yields, while meso - 49 was converted to the single mono-

acetate enantiomer ( R,S,R,S ) - 51 via efﬁ cient desymmetrization (see equation 12,

Scheme 12.4 ) [4b, 4e] . Since these mono - and bis - gem - diﬂ uorocyclopropanes have two

hydroxymethyl groups to modify, a variety of compounds can be prepared using them as

building blocks [4, 22] .

de Meijere and co - workers reported the synthesis of enantiopure 7,7 - diﬂ uorodispiro

[2.0.2.1]heptylmethanols ( 54 ) via diﬂ uorocyclopropanation of methylenespirobiscyclo-

propane (1 S ,3 R ) - 53 ( > 99% ee), which was obtained by lipase - catalyzed enantiomer -

speciﬁ c transesteriﬁ cation of ( ± ) - 52 , followed by separation of two diastereomers (see

equation 13, Scheme 12.4 ) [30] .

Scheme 12.4 Preparation of optically active gem - diﬂ uorocyclopropanes using enzymatic

resolution.

326 Fluorine in Medicinal Chemistry and Chemical Biology

12.5 Biologically Active gem - Diﬂ uorocyclopropanes:

Anthracene – gem - Diﬂ uorocyclopropane Hybrids as DNA Cleavage

Agents Switched on by Photoirradiation

As described above, gem - diﬂ uorocyclopropane analogues of structurally complex

biologically active natural products reported to date have not shown enhanced activities

[8, 14] , while rather simple synthetic gem - diﬂ uorocyclopropanes have shown interesting

biological activities [15, 16, 18 – 22] . We recently reported the synthesis of gem -

diﬂ uorocyclopropanes that showed a strong DNA - cleaving property switched on by

photoirradiation [22] .

During the course of determining the stereochemistry of 1,3 - bishydroxymethyl - 2,2 -

diﬂ uorocyclopropane on the basis of CD (circular dischroism) spectroscopic analysis of

the corresponding 9 - anthracenecarboxylic acid diester 55 , we recognized the interesting

fact that diester 55 was so unstable in dichloromethane solution under sunlight that it

formed unidentiﬁ ed complex compounds by opening its gem - diﬂ uorocyclopropane ring

[4g] . We also found that, similar to this mono - diﬂ uorocyclopropane 55 , the anthracenecar-

boxylic acid diester of bis - gem - diﬂ uorocyclopropane was rapidly decomposed by expo-

sure to sunlight [4c] . It has been shown that anthracene or anthraquinone derivatives serve

as potent DNA - cleavage agents [31 – 34] . For example, Schuster and co - workers reported

that anthraquinonecarboxamide caused GG - selective cleavage of duplex DNA [32] , while

Kumar and Punzalan found that anthracene - substituted alkylamine derivatives possessed

potent DNA - cleaving activity [33] . Toshima et al. reported that quinoxaline - carbohydrate

hybrids acted as GG - selective DNA - cleaving or DNA - binding agents, depending on the

structure of the sugar moiety [34] . From these results, we anticipated that our anthracene -

gem - diﬂ uorocyclopropane hybrids would possess DNA - cleaving activity on photoirradia-

tion (see Figure 12.9 ). It has been shown that decomposition of a gem - diﬂ uorocyclopropane

ring proceeds via a radical species [35] , which may cause DNA - damage [36] . We

prepared optically active anthracenecarbonyl derivatives of gem - diﬂ uorocyclopropane,

19, 55 , and 56 , and evaluated their DNA - cleaving activities. As shown in Figure 12.9 ,

( S,S ) - 55 (lane II) and ( S,S ) - 56 (lane IV) showed only weak DNA - cleaving activities,

while much stronger activity was exhibited by anthracenecarboxamidomethyl - gem -

diﬂ uorocyclopropane 19 . In the presence of 100 µ M of ( S,S ) - 19 , which corresponds to 1.3

molar equivalents to a DNA base pair, DNA was cleaved by photoirradiation and super-

coiled φ X174 DNA (Form I) was converted to the relaxed form (Form II) (see Figure 12.9 ,

lane V). No signiﬁ cant difference in activity was observed between the enantiomers of 19 ,

i.e., ( S,S ) - 19 (lane V) and ( R,R ) - 19 (lane IV). Although nonﬂ uorinated cyclopropane

counterpart ( S,S ) - 57 also caused substantial DNA - cleavage (lane VI), the activity of

gem - diﬂ uorocyclopropane ( S,S ) - 19 was obviously superior to that of ( S,S ) - 57 at the same

concentration. However, no sequence speciﬁ city was observed in these DNA - cleavage

experiments.

We also found an interesting difference in DNA - cleaving activity between the enan-

tiomers of aminomethyl - gem - diﬂ uorocyclopropylmethyl anthracenecarboxylate, ( S,S ) - 18

and ( R,R ) - 18 . As Figure 12.10 shows, ( R,R ) - 18 exhibited ∼ 10 - fold stronger DNA - cleaving

activity than ( S,S ) - 18 for supercoiled φ X174 DNA. The result clearly indicates that the

gem-Diﬂ uorocyclopropanes as Key Building Blocks for Novel Biologically Active Molecules 327

chirality of the gem - diﬂ uorocyclopropane moiety controls the DNA - cleaving activity of

these anthracene – gem - diﬂ uorocyclopropane hybrids.

These results possibly suggest that the DNA - cleavage by these anthracene – gem -

diﬂ uorocyclopropane hybrids is caused mainly by intercalation or minor - groove binding

of the anthracenyl group to DNA and the gem - diﬂ uorocyclopropane component plays an

important role in determining the DNA - binding mode. Although we found no differences

in sequence speciﬁ city between ( S,S ) - 18 and ( R,R ) - 18 , we hypothesize that the terminal

amino group may bind to the phosphate moiety of DNA, and then direct the intercalation

Figure 12.9 Photocleavage of supercoiled φ X174 plasmid DNA by cyclopropane derivatives.

Lanes: c, control; I, ethyl 9 - anthracenecarboxylate; II, (S,S) - 55 ; III, (S,S) - 56 ; IV, (R,R) - 19 ;

V, (S,S) - 19 ; VI, (S,S) - 57 . Reaction buffer: 20 mM sodium phosphate pH 7.0 (20% DMSO)

Samples were irradiated at 25 ° C for 45 min at a distance of 6 cm using a xenon lamp with

a polystyrene ﬁ lter (365 nm). Electrophoresis was run using 0.8% agarose gel with TAE

buffer at 100 V for 40 min. The gel was stained with EtBr and visualized by UV - B lamp

(transilluminator).