Bioorganic & Medicinal Chemistry 22 (2014) 3922–3930

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Bioorganic & Medicinal Chemistry

journal homepage: www.elsevier .com/locate /bmc

Synthesis and fungicidal activity of quinolin-6-yloxyacetamides,a novel class of tubulin polymerization inhibitors

http://dx.doi.org/10.1016/j.bmc.2014.06.0150968-0896/� 2014 Elsevier Ltd. All rights reserved.

⇑ Corresponding author. Tel.: +41 62 866 0224; fax: +41 62 866 0860.E-mail address: clemens.lamberth@syngenta.com (C. Lamberth).

Clemens Lamberth a,⇑, Fiona Murphy Kessabi a, Renaud Beaudegnies a, Laura Quaranta a, Stephan Trah a,Guillaume Berthon a, Fredrik Cederbaum a, Gertrud Knauf-Beiter b, Valeria Grasso b, Stephane Bieri b,Andy Corran c, Urvashi Thacker c

a Syngenta Crop Protection AG, Research Chemistry, Schaffhauserstr. 101, CH-4332 Stein, Switzerlandb Syngenta Crop Protection AG, Research Biology, Schaffhauserstr. 101, CH-4332 Stein, Switzerlandc Syngenta Ltd, Jealott’s Hill International Research Centre, Bracknell, Berkshire RG42 6EY, United Kingdom

a r t i c l e i n f o

Article history:Received 7 April 2014Revised 3 June 2014Accepted 6 June 2014Available online 17 June 2014

Keywords:TubulinQuinolineHeterocycleFungicideCrop protection

a b s t r a c t

A novel class of experimental fungicides has been discovered, which consists of special quinolin-6-ylo-xyacetamides. They are highly active against important phytopathogens, such as Phytophthora infestans(potato and tomato late blight), Mycosphaerella graminicola (wheat leaf blotch) and Uncinula necator(grape powdery mildew). Their fungicidal activity is due to their ability to inhibit fungal tubulin polymer-ization, leading to microtubule destabilization. An efficient synthesis route has been worked out, whichallows the diverse substitution of four identified key positions across the molecular scaffold.

� 2014 Elsevier Ltd. All rights reserved.

1. Introduction

Microtubules are hollow cylindrical tubes found in all eukary-otic cell types. These essential cytoskeletal protein polymers playa pivotal role in maintaining the growth, shape, division, motilityand functioning of the cell. Microtubules are key components ofthe mitotic spindle, which enables the segregation of chromo-somes during the process of mitosis. They are built by polymeriza-tion of the two globular protein subunits a- and b-tubulin, whichfirst combine to a,b-heterodimers. Interference with the microtu-bule homeostasis by disrupting the dynamic equilibrium betweenthe depolymerization of microtubules into tubulin or, inversely,the assembly of tubulin into microtubules leads to arrested celldivision and consequently to apoptosis.1 Especially this latter pos-sibility, the inhibition of the tubulin polymerization, also calledmicrotubule destabilization, has widely impacted the chemothera-peutic treatment of human cancer diseases2 as well as the protec-tion of plants against fungal diseases.3 A group of special naturalproducts, such as the vinca alkaloids,4 for example vinblastine,5

vincristine,6 vinorelbine7 and vindesine,8 but also colchicine9 and

combretastatin10 are a well-established class of anti-cancer agents.On the other hand tubulin polymerization inhibitors such as themethyl benzimidazole carbamates (MBC’s),11 for example beno-myl,12 carbendazim,13 thiabendazole14 and fuberidazole,15 are suc-cessfully applied as agrochemical fungicides.

Although much effort is currently put into the search for noveltubulin polymerization inhibitors, there is clearly a lack of addi-tional, independent subclasses. In oncology, many of the identifiedactive compounds depend on scaffolds based on the well-known3,4,5-trimethoxyphenyl ring of colchicine and combretastatin.16–25

In crop protection, the introduction of the MBC’s as very first sys-temic active fungicide class in the 1960s revolutionized the agro-chemical market, but in the following decades with zoxamide26

and ethaboxam27 only two further tubulin polymerization inhibi-tors have been developed. Therefore there is definitely a need foractive ingredients with completely novel structural entities.

In this paper we present quinolin-6-yloxyacetamides as a novelclass of tubulin polymerization inhibitors.28–30 Their discovery wasclearly fertilized by cross-indication screening, because the glyoxy-lic acid acetal derivative 1, belonging to a herbicide project, wasidentified as a fungicide hit owing to some weak, but interestingsignals in the greenhouse (Fig. 1). The thorough optimization ofthis lead compound led to quinolin-6-yloxyacetamides such as 2,which shows powerful fungicidal activity in field trials.

N

ONH

O

S

NOO

NH

O

O

Cl

Cl

1 2

initial lead structurefrom herbicide project

typical example of fungicidallyactive quinolin-6-yloxyacetamides

Figure 1. Typical quinolin-6-yloxyacetamide 2 and its original lead compound 1.

C. Lamberth et al. / Bioorg. Med. Chem. 22 (2014) 3922–3930 3923

2. Results and discussion

2.1. Chemistry

3-Bromo-8-methylquinolin-6-ol (7) is a key building block inthe synthesis of the fungicidally active quinolin-6-yloxyacetamides9 and 12 (Scheme 1). We have recently described the efficient two-step preparation of this compound via Skraup-type cyclizationfrom either 2-methyl-4-nitroaniline (3) or 4-methoxy-2-methy-laniline (5) with 2,2,3-tribromopropanal.31 The hydroxyl functionin position 6 of the trisubstituted quinoline 7 can be easily alkyl-ated with a-halocarboxylates, for example, leading to the quino-lin-6-yloxyacetates 8 and 10 bearing either an ethyl or athiomethyl group at the a-carbon atom of the ester. These twofunctionalities played the biggest role during our optimization,but also several further substituents have been introduced bychoice of the appropriate a-chlorocarboxylate (Table 4). It is note-worthy, that within this alkylation reaction, completely differentfunctional groups can be produced under the same reaction condi-tions, as the hydroxyl function of 7 has been transformed into anether (?8) as well as into a O,S-acetal (?10). The saponificationof the ester function in 8 and 10 to the corresponding carboxylicacids and their further conversion with tert-butylamine and prop-argylamine, respectively, under peptide coupling conditions finallyled to the fungicidally active amides 9 and 12.

The methyl ester 10, which is a key intermediate in the synthesisof the fungicidally active amide 12, is perfectly suited for the trans-formation into derivatives with a different quinoline substitutionpattern by exchange of the bromine atom. During our optimizationof this compound class it turned out that quinolin-6-yloxyaceta-mides with a carbon-linked substituent in position 3 have beenespecially interesting. Scheme 2 describes the palladium-catalyzedreplacement of the bromo function by four different carbon substit-uents, an alkyl, an alkenyl, an alkynyl and a cycloalkyl group. Thevinyl derivative 13, which can be converted into the final product

NH2

NO2

NH2

O

N

Br NO2

N

Br O

N

Br O

SOH

O

N

Br O

SNH

O

4

6

1. Fe, HCl2. H3PO4, H2O

CH3 CH3

77 % 67 %

45 %

HBr

84 %

CH2 BrCBr2CHOAcOH

CH2 BrCBr2CHOAcOH

1112

3

5

NaOHHOAt, EDCI,tBuNH2,NEt3

67 % 100 %

Scheme 1. Synthesis of the fungicidally activ

14, is obtained from bromoquinoline 10 by a palladium-catalyzedStille reaction with vinyltributyltin.32 Another possibility for thesynthesis of 13 is the Sonogashira coupling of trimethylsilylacety-lene33 and 10 to the 3-ethynylquinoline 15 and the subsequent par-tial reduction of the C–C triple bond by catalytic hydrogenationwith Lindlar catalyst. In addition, the ester 15 can also convertedinto the fungicidally active amide 2. The 3-methylquinolin-6-yloxyacetate 16, which delivers the corresponding amide 17, is obtainedby Suzuki–Miyaura coupling with trimethylboroxine.34 Finally, thereplacement of the bromo substituent of 10 by cyclopropyl wasachieved by Suzuki–Miyaura coupling with cyclopropylboronicacid,35 leading to the ester 18 which delivers the target compound19 upon amidation. These transformations confirm the huge scopeof palladium-catalyzed cross-coupling reactions of halogenatedquinolines, a field which has been reviewed recently.36

2.2. Mode of action

The quinolin-6-yloxyacetamides 2 and 12 were submitted to apolymerization assay on pure porcine tubulin to check if theseexperimental fungicides are disrupters of the microtubule dynam-ics. They were compared against colchicine, which is a knowntubulin polymerization inhibitor and for which a strong effect onthe OD340 value could be detected at 4 lg ml�1. Also the two quin-olin-6-yloxyacetamides 2 and 12 showed in two different repli-cates an inhibitory effect on the microtubule formation, whichwas clearly different from the solvent control (Fig. 2).

2.3. Structure–activity relationships

During our derivatisation of this novel class of tubulin polymer-ization inhibitors we identified four key positions in the molecularscaffold of quinolin-6-yloxyacetamides, which have to bear theright substituents for optimum fungicidal activity. Tables 1–5describe the effect of replacement of the quinoline core by otherrings and of those four important quinoline substituents on theefficacy against the Oomycetes disease Phytophthora infestans(potato and tomato late blight) as well as against Mycosphaerellagraminicola (wheat leaf blotch) and Uncinula necator (grape pow-dery mildew) which are both from the family of Ascomycetes.

2.3.1. Influence of the cyclic core on the fungicidal activityOne of the most important breakthroughs during our derivati-

sation was the discovery, that the 3-bromoquinolin-6-yl scaffold(Table 1, entry 3) is a big improvement compared to the dihalogen-ated benzene ring of the initial lead compound 1 (entry 1). Conse-

N

Br OH

N

Br O

SO

O

N

Br OO

O

N

Br ONH

O

CH CCH2NH2

7

10

8

9

ClCH(SMe)CO2Me,K2CO3

BrCH(Et)CO2Me,K2CO3

1. LiOH2. HOAt, EDCI,

91 %

43 % overtwo steps

NEt3,71 %

e quinolin-6-yloxy acetamides 9 and 12.

N

Br O

SO

O

N

O

SO

O

N

O

SO

O

N

O

SO

O

N

O

SO

O

CH CSiMe3

N

O

SNH

ON

O

N

O

SNH

OO

N

N

O

SNH

O

Cl

N

O

SNH

O

O

MeOCH2C CC(Me)2NH2

1015

13

16

18

(MeBO)3,Pd(PPh3)4K2CO3

c-PrB(OH)2,Pd(PPh3)4K3PO4

H2,Pd/Lindlarquinoline

H2C=CHSnBu3,

Pd(PPh3)4

PdCl2(PPh3)2,CuI, iPr2NH

2. K2CO3

2

14

19

17

1.

1. NaOH2. HOAt, TBTU, NEt3

MeOCH2C(Me)(CN)NH2

54 % overtwo steps

49 % overtwo steps

47 %

1. NaOH2. HOAt, EDCI, NEt3, MeON=CHC(Me)2NH2

56 % overtwo steps

51 %

1. NaOH2. HOAt, TBTU, NEt3,

25 %

1. NaOH2. HOAt, EDCI, NEt3 H2C=C(Cl)CH2NH2

56 % overtwo steps

82 % overtwo steps

61 %

Scheme 2. Transformation of the 3-bromoquinoline derivative 10 into different 3-carba-substituted quinolin-6-yloxyacetamides.

Figure 2. Degree of polymerization of pure porcine tubulin in the presence ofcolchicine (4 lg ml�1), 2 (1 lg ml�1) and 12 (3 lg ml�1) in function of time (valuespresented are a mean of two replicates).

3924 C. Lamberth et al. / Bioorg. Med. Chem. 22 (2014) 3922–3930

quently, much effort was invested in the fine-tuning of thissuccessful heterobicyclic core. However, shifting the ring nitrogento the other ring (entry 4), replacing it by a carbon atom (entry 5)or adding a second ring nitrogen to the bicycle (entry 6) clearlydecreased the fungicidal potency. Also the replacement of the bro-moquinoline of entry 3 by a bromobenzothiophene (entry 7) didnot result in sufficient biological activity.

2.3.2. Influence of the substituent in quinoline position 3 on thefungicidal activity

The excellent results of several differently 3-substituted quino-lines in Table 2 demonstrate the huge scope of variability in thisposition. Relatively soon after the discovery of the quinoline scaf-fold we identified the higher halogens bromine (entry 3) andiodine (entry 4) as highly active substituents in this position. Thecorresponding chloro (entry 2) derivative fails completely againstPhytophthora infestans. The only other substituent which delivers

at least equal results than bromo and iodo is the ethynyl group(entry 8). This is rather surprising, as the other C2 groups withreduced degree of unsaturation, ethyl (entry 6) and vinyl (entry7) show clearly weaker fungicidal potency. Also methoxy (entry10) cannot compete with bromo, iodo and ethynyl. Most of thesecarbon- and oxygen-linked substituents have been introduced byderivatization of the 3-bromoquinoline building block 10(Scheme 2).

2.3.3. Influence of the substituent on quinoline position 8 onthe fungicidal activity

Ring carbon 8 is the third quinoline ring position, which wasidentified besides positions 3 and 6 as crucial to be substitutedwith the optimum functional group for delivery of the best possiblefungicidal activity (Table 3). The substituent in this position couldbe broadly varied by choice of the appropriate aniline startingmaterial as shown in Scheme 1.28 It seems that a methyl groupin this position (entry 6) delivers the highest fungicidal activity,followed by the 8-unsubstituted analog (entry 1) and the chlori-nated (entry 3) and brominated (entry 4) derivatives. Interestingly8-iodo substituted quinolin-6-yloxyacetamide (entry 5) is clearlyweaker than its bromo analog (entry 4), whereas in ring position3 (Table 2), iodo was more or less equal to bromo. Much efforthas been undertaken to increase the fungicidal potential of the 8-methyl substituted compounds by small structural modifications.However, the elongation of the methyl group to the next higherhomolog ethyl (entry 7) as well as the replacement of hydrogenatoms of the methyl group by fluorine atoms (entries 8 and 9)led to a clear drop in fungicidal activity.

2.3.4. Influence of the substituent in the acetic acid moiety onthe fungicidal activity

The completely missing fungicidal activity of a-unsubstitutedquinolin-6-yloxyacetamides (entry 1) demonstrates already theimportance of the right substituent in this position of the aceticacid moiety (Table 4). In our experience, two-atom side chains,especially the ethyl group (entry 2) and the methylthio group(entry 6) deliver the best results. Already small modifications lead

Table 1Influence of the cyclic core on the fungicidal activitya

Entry R Phytophthorainfestans(potato andtomato lateblight)

Mycosphaerellagraminicola(wheat leafblotch)

Uncinulanecator(grapepowderymildew)

1 67 15 30

2 13 0 0

3 95 95 100

4 47 0 0

5 10 0 88

6 8 0 0

7 57 0 59

a Results are given in % activity at 60 ppm.

Table 3Influence of the substituent in quinoline position 8 on the fungicidal activitya

Entry R Phytophthorainfestans (potatoand tomato lateblight)

Mycosphaerellagraminicola(wheat leafblotch)

Uncinulanecator (grapepowderymildew)

1 H 95 95 1002 F 83 87 1003 Cl 96 93 1004 Br 90 100 1005 I 0 79 36 CH3 100 100 1007 CH2CH3 20 82 958 CHF2 0 100 289 CF3 8 0 010 CN 3 93 9811 C„CH 80 98 10012 SCH3 0 0 0

a Results are given in % activity at 60 ppm.

Table 4Influence of the substituent in the acetic acid moiety on the fungicidal activitya

Entry R1 R2 Phytophthorainfestans(potato andtomato lateblight)

Mycosphaerellagraminicola(wheat leafblotch)

Uncinulanecator(grapepowderymildew)

1 H H 0 0 02 CH2CH3 H 87 90 1003 CH2CH2F H 61 0 1004 OCH3 H 0 74 1005 OCH2CH2OCH3 H 46 11 06 SCH3 H 95 95 1007 SCH2CH3 H 14 0 98 SPh H 1 0 09 Pyrazol-1-yl H 60 17 9010 CH2CH3 F 0 4 011 SCH3 CH3 0 14 012 –CH2CH2– 1 0 0

a Results are given in % activity at 60 ppm.

Table 2Influence of the substituent in quinoline position 3 on the fungicidal activitya

Entry R Phytophthorainfestans (potatoand tomato lateblight)

Mycosphaerellagraminicola(wheat leafblotch)

Uncinulanecator (grapepowderymildew)

1 H 97 100 752 Cl 0 94 1003 Br 100 100 1004 I 99 100 1005 CH3 75 80 1006 CH2CH3 63 90 997 CH@CH2 65 25 168 C„CH 100 100 1009 C„CCH3 0 0 7610 OCH3 13 75 100

a Results are given in % activity at 60 ppm.

C. Lamberth et al. / Bioorg. Med. Chem. 22 (2014) 3922–3930 3925

to dramatic differences in the fungicidal activity, for example thereplacement of the sulfur atom in the methylthio group by oxygen(entry 4) and the elongation of the methylthio group by oneadditional carbon atom (entry 7). The attempt to enhance theactivity of the two best substituents ethyl and methylthio by intro-duction of an additional substituent, leading to a quarternization ofthe a-acetic acid carbon was not successful at all (entries 10 and

11). Also another quarternary derivative, in which the a-carbonis part of a cyclopropyl ring, was devoid of any fungicidal activity(entry 12).

2.3.5. Influence of the amine moiety on the fungicidal activityThe abundant availability of amines enabled us to check the full

scope of this terminal functional group of our quinolin-6-yloxyac-etamides. Table 5 compares different primary, secondary and ter-tiary amides of the same acid building block. Hereby someinteresting trends became visible. A NH2 group linked to the acid(primary amide, entry 1) is devoid of any useful fungicidal activity.The best results regarding level of activity and broadness of spec-trum have been achieved with the secondary amides (entries 7–9), in which the quarternary carbon atom next to the amine nitro-gen is substituted by either three methyl groups (entry 7), twomethyl groups and another carbon substituent (entry 8) or one

Table 5Influence of the amine moiety on the fungicidal activitya

Entry R1 R2 Phytophthora infestans(potato and tomato late blight)

Mycosphaerella graminicola(wheat leaf blotch)

Uncinula necator(grape powdery mildew)

1 H H 0 46 02 CH2CH(CH3)2 H 0 80 1003 CH2CH@CH2 H 0 90 1004 CH2C(CH3)@CH2 H 0 90 1005 CH(CH3)CH2CH3 H 0 80 986 CH(CH3)CH2CF3 H 11 99 987 C(CH3)3 H 100 100 1008 C(CH3)2C„CH H 100 100 1009 C(CH3)(CN)CH2OCH3 H 100 100 10010 C(CH3)2(4-ClPh) H 21 0 1011 C(CH3)3 CH3 1 72 7612 AC(@O)OCH2C(Me)2- 11 0 63

a Results are given in % activity at 60 ppm.

3926 C. Lamberth et al. / Bioorg. Med. Chem. 22 (2014) 3922–3930

methyl group and two other carbon functional groups (entry 9).This impressive level of activity drops immediately, if one of thesethree carbon substituents at the a-amine carbon exceeds a certainsize, such as the p-chlorophenyl ring in entry 10. Also the introduc-tion of a second alkyl group into the amide nitrogen (tertiaryamide, entry 11) or the incorporation of the amide nitrogen intoan oxazolidinone ring (entry 12) decreased the fungicidal activity.The fact that in general, secondary amides with a CH2 group(entries 2–4) or a CH(Me) group (entries 5 and 6) next to the aminenitrogen selectively lost the activity against the Oomycetes patho-gen P. infestans, but kept a high level of efficacy against the Asco-mycetes diseases M. graminicola and U. necator was a fascinatingdiscovery.

3. Conclusions

Novel quinolin-6-yloxyacetamides have been discovered as anew class of fungicidally active compounds, which are able to con-trol economically important plant diseases, such as P. infestans, M.graminicola and U. necator. Their mode of action is the inhibition oftubulin polymerization, leading to microtubule destabilization.They can be prepared starting from different 4-methoxy- or 4-nitroanilines, which deliver in a Skraup-type transformation with2,2,3-tribromopropanal 3-bromoquinolin-6-ols. These key inter-mediates can be converted in only few steps into a high numberof quinolin-6-yloxyacetamides with different substituents in thequinoline core, the acetic acid part and the amine moiety.

A structure–activity relationship study revealed the molecularrequirements for the best fungicidal activity. We first discoveredthe superiority of the quinoline scaffold compared to several othermono- and bicyclic analogs and then identified four key positionswhich have to be specifically substituted to reach optimum effi-cacy. Bromo, iodo and ethynyl are by far the best substituents inquinoline position 3, whereas the ring position 8 should bear ahydrogen atom or a methyl group for optimum results and an ethylor methylthio group linked to the a-carbon atom of the acetic acidmoiety. This means, that completely different substituents arerequired in these three positions for the best fungicidal activity.Furthermore the amine side chain plays an important role for thefungicidal activity. The highest level of activity was achieved bysecondary amides, in which the amino function is linked to a quar-ternary carbon atom. Interestingly, the activity against Oomycetes

diseases, such as P. infestans, can be completely surppressed, whenthe a-carbon atom of the amine is unsubstituted.

4. Experimental section

4.1. Chemistry

All new compounds were characterized by standard spectro-scopical methods. 1H NMR spectra were recorded on a Varian Unity400 spectrometer at 400 MHz using CDCl3 as solvent and tetra-methylsilane as internal standard. Chemical shifts are reported inppm downfield from the standard (d = 0.00), coupling constantsin Hz. LC-MS spectra were determined using the following appara-tus: ACQUITY UPLC from Waters, Phenomenex Gemini C18, 3 mmparticle size, 110 Angström, 30 � 3 mm column, 1.7 ml/min, 60 �C,H2O + 0.05% HCOOH (95%)/CH3CN/MeOH 4:1 + 0.04% HCOOH(5%)—2 min—CH3CN/MeOH 4:1 + 0.04% HCOOH (5%)—0.8 min;ACQUITY SQD Mass Spectrometer from Waters, ionization method:electrospray (ESI), Polarity: positive ions, Capillary (kV) 3.00, Cone(V) 20.00, Extractor (V) 3.00, Source Temperature (�C) 150, Desolv-ation Temperature (�C) 400, Cone Gas Flow (L/Hr) 60, DesolvationGas Flow (L/Hr) 700. Analytical thin-layer chromatography (TLC)was performed using silica gel 60 F524 precoated plates. Prepara-tive flash chromatography was performed using silica gel 60(40–63 lm, E. Merck). Unless otherwise stated, all reactions werecarried out under anhydrous conditions in an inert atmosphere(nitrogen or argon) with dry solvents.

4.1.1. 3-Bromo-8-methyl-6-nitroquinoline(4)2,2,3-Tribromopropanal28 (29.4 g, 0.1 mol) was slowly added to

a suspension of 2-methyl-4-nitroaniline (3, 15,2 g, 0.1 mol) in200 ml of glacial acetic acid. The reaction mixture was heated to110 �C for 1 h, then cooled to room temperature and filtered. Theremaining solid was washed with diethyl ether, then suspendedin water and treated with a saturated aqueous sodium bicarbonatesolution until pH 9 was reached. The suspension was transferred toa separatory funnel and extracted with ethyl acetate, the organicphase was dried over magnesium sulfate and evaporated underreduced pressure. The residue was purified by chromatographyon silica gel, using ethyl acetate and heptane as eluents to obtain3-bromo-8-methyl-6-nitroquinoline (4, 20.7 g, 77 mmol, 77%). 1HNMR (CDCl3): d = 2.86 (s, 3H), 8.35 (d, 1H, J = 2.1 Hz), 8.47 (d, 1H,

C. Lamberth et al. / Bioorg. Med. Chem. 22 (2014) 3922–3930 3927

J = 2.0 Hz), 8.54 (d, 1H, J = 2.2 Hz), 9.06 (d, 1H, J = 2.1 Hz). LC-MS:Rt = 1.94 min; MS: m/z = 268 [M]+, 269 [M+1]+.

4.1.2. 3-Bromo-6-methoxy-8-methylquinoline (6)2,2,3-Tribromopropanal (50.0 g, 0.18 mol) was slowly added to a

suspension of 4-methoxy-2-methylaniline (5, 25.0 g, 0.18 mol) in300 ml of glacial acetic acid. The reaction mixture was stirred for6 h at room temperature, then diluted with ethyl acetate and washedwith water, brine and 2 N sodium hydroxide solution, subsequentlydried over magnesium sulfate and evaporated under reduced pres-sure. The residue was purified by chromatography on silica gel, usingethyl acetate and heptane as eluents to yield 3-bromo-6-methoxy-8-methylquinoline (6, 20.5 g, 81 mmol, 45%). 1H NMR (CDCl3):d = 2.73 (s, 3H), 3.90 (s, 3H), 6.82 (d, 1H, J = 2.0 Hz), 7.21 (d, 1H,J = 2.1 Hz), 8.17 (d, 1H, J = 1.9 Hz), 8.76 (d, 1H, J = 2.0 Hz). LC-MS:Rt = 1.99 min; MS: m/z = 254 [M]+, 255 [M+1]+.

4.1.3. 3-Bromo-8-methylquinolin-6-ol (7)From 4: reduced iron powder (15 g, 0.27 mol) was added in por-

tions to a suspension of 3-bromo-8-methyl-6-nitroquinoline (4,20.6 g, 77 mmol) in a mixture of 400 ml of ethanol and 2 ml of37% aqueous hydrochloric acid at room temperature. The reactionmixture was heated to reflux for 2 h, during which the color of thesuspension changed from grey–yellow to red–brown. The reactionmixture was cooled to 40 �C, filtered through Celite, the filtratewas diluted with ethanol, treated with silica gel and concentratedunder reduced pressure. The residue was purified by chromatogra-phy on silica gel, using ethyl acetate and dichloromethane as eluentsto deliver 6-amino-3-bromo-8-methylquinoline. This intermediatewas suspended in a mixture of 125 ml of 85% aqueous phosphoricacid and 12 ml of water and heated in a tantalum pressure vesselto 180 �C for 72 h. Subsequently, the mixture was cooled to roomtemperature and poured on water. 30% Aqueous sodium hydroxidewas added to this solution until pH 2–4 was reached. The precipitateformed was filtered, washed with cold water and dried to give 3-bromo-8-methylquinolin-6-ol (7, 12.3 g, 52 mmol, 67%). 1H NMR(DMSO-d6): d = 2.64 (s, 3H), 6.99 (d, 1H, J = 2.1 Hz), 7.22 (d, 1H,J = 2.1 Hz), 8.47 (d, 1H, J = 2.2 Hz), 8.69 (d, 1H, J = 2.3 Hz), 10.13 (s,1H). LC-MS: Rt = 1.78 min; MS: m/z = 238 [M]+, 239 [M+1]+.

From 6: a mixture of 3-bromo-6-methoxy-8-methylquinoline(6, 14.0 g, 55 mmol) in 250 ml of 48% hydrobromic acid was slowlyheated to 110 �C and kept at this temperature for 20 h. Subse-quently, the reaction mixture was cooled to room temperatureand filtered. The residue was washed with water, taken up in sat-urated aqueous sodium bicarbonate solution and filtered again.The remainder was washed with water and dried in high vacuumto afford 3-bromo-8-methylquinolin-6-ol (7, 11.1 g, 47 mmol,84%). 1H NMR and LC-MS were identical to those obtained from 4.

4.1.4. 2-(3-Bromo-8-methylquinolin-6-yloxy)-butyric acidmethyl ester (8)

Potassium carbonate (8.7 g, 63 mmol) was added to a solutionof 3-bromo-8-methylquinolin-6-ol (7, 5.0 g, 21 mmol) in 70 ml ofN,N-dimethylformamide. After the addition of methyl 2-bromobu-tyrate (7.1 g, 38 mmol), the reaction mixture was stirred for 16 h atroom temperature. Subsequently the mixture was filtered and thefiltrate was diluted with ethyl acetate. This organic phase waswashed with water and brine, dried over magnesium sulfate andevaporated under reduced pressure. The residue was purified bychromatography on silica gel, using ethyl acetate and cyclohexaneas eluents to yield 2-(3-bromo-8-methylquinolin-6-yloxy)-butyricacid methyl ester (8, 6.5 g, 19 mmol, 91%). 1H NMR (CDCl3):d = 1.04 (t, 3H, J = 7.3 Hz), 1.99 (q, 2H, J = 7.1 Hz), 2.68 (s, 3H),3.70 (s, 3H), 4.63 (t, 1H, J = 7.7 Hz), 6.65 (d, 1H, J = 2.3 Hz), 7.22(d, 1H, J = 2.2 Hz), 8.06 (d, 1H, J = 2.1 Hz), 8.68 (d, 1H, J = 2.1 Hz).LC-MS: Rt = 1.12 min; MS: m/z = 340 [M+1]+, 341 [M+2]+.

4.1.5. 2-(3-Bromo-8-methylquinolin-6-yloxy)-N-prop-2-ynyl-butyramide (9)

Lithium hydroxide hydrate (0.2 g, 5.3 mmol) was added to a solu-tion of 2-(3-bromo-8-methylquinolin-6-yloxy)-butyric acid methylester (8, 1.5 g, 4.4 mmol) in a mixture of 11 ml of tetrahydrofuranand 11 ml of water at 0 �C The reaction mixture was stirred for 2 hat 0 �C and then warmed to room temperature. The tetrahydrofuranwas evaporated under reduced pressure and 1 N hydrochloric acidwas added to the remaining mixture until pH 1 was reached. The pre-cipitate formed was filtered and dried in a vaccum oven at 50 �C todeliver 2-(3-bromo-8-methyl-quinolin-6-yloxy)-butyric acid. Thisintermediate was dissolved in 30 ml of N,N-dimethylformamideand triethylamine (1.5 g, 15 mmol), 1-hydroxy-7-azabenzotriazole(0.9 g, 6.5 mmol), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide(1.2 g, 6.5 mmol) and propargylamine (0.4 g, 6.5 mmol) were addedconsecutively. The reaction mixture was stirred for 16 h at roomtemperature, then diluted with ethyl acetate and extracted withbrine, saturated aqueous sodium bicarbonate solution and water.The organic layer was dried over magnesium sulfate and evaporatedunder reduced pressure, the residue was crystallized with t-butylmethyl ether to deliver 2-(3-bromo-8-methylquinolin-6-yloxy)-N-prop-2-ynylbutryramide (9, 0.7 g, 1.9 mmol, 43%). Mp 174–178 �C(%). 1H NMR (CDCl3): d = 1.00 (t, 3H, J = 7.2 Hz), 1.92–2.03 (m, 2H),2.11 (d, 1H, J = 2.4 Hz), 2.78 (s, 3H), 3.93 (dd, 1 H, J = 2.3 Hz,11.0 Hz),4.06 (dd, 1H, J = 2.4 Hz, 10.8 Hz), 4.59 (t, 1H, J = 7.5 Hz), 6.45 (br s,1H), 6.76 (d, 1H, J = 2.0 Hz), 7.22 (d, 1H, J = 2.1 Hz), 8.09 (d, 1H,J = 2.2 Hz), 8.72 (d, 1H, J = 2.1 Hz). LC-MS: Rt = 0.97 min; MS:m/z = 363 [M+1]+, 364 [M+2]+.

4.1.6. 2-(3-Bromo-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (10)

Methyl 2-chloro-2-(methylsulfanyl)acetate (11 g, 73 mmol) andmilled potassium carbonate (42.5 g, 0.3 mol) were consecutivelyadded to a suspension of 3-bromo-8-methylquinolin-6-ol (7,14.5 g, 61 mmol) in 200 ml of N,N-dimethylformamide. The reac-tion mixture was stirred for 2 h at 60 �C, then cooled to room tem-perature and diluted with ethyl acetate. The organic layer waswashed with water, dried over magnesium sulfate and evaporatedunder reduced pressure. The residue was purified by chromatogra-phy on silica gel, using ethyl acetate and hexane as eluents todeliver 2-(3-bromo-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (10, 15.5 g, 43 mmol, 71%). 1H NMR(CDCl3): d = 2.23 (s, 3H), 2.74 (s, 3H), 3.87 (s, 3H), 5.72 (s, 1H),6.96 (d, 1H, J = 2.2 Hz), 7.36 (d, 1H, J = 2.1 Hz), 8.18 (d, 1H,J = 2.0 Hz), 8.80 (d, 1H, J = 2.1 Hz). LC-MS: Rt = 2.04 min; MS:m/z = 358 [M+1]+, 359 [M+2]+.

4.1.7. 2-(3-Bromo-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid (11)

2 N sodium hydroxide (21 ml, 42 mmol) was added to a suspen-sion of 2-(3-bromo-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (10, 10 g, 28 mmol) in 65 ml of ethanol.The reaction mixture was stirred for 1 h at room temperature, thencooled to 0 �C and acidified with 2 N hydrochloric acid until pH 2was reached. The resulting precipitate was filtered and dried in adessicator to obtain 2-(3-bromo-8-methylquinoline-6-yloxy)-2-methylsulfanyl-acetic acid (11, 9.6 g, 28 mmol, 100%). 1H NMR(DMSO-d6): d = 2.17 (s, 3H), 2.68 (s, 3H), 6.06 (s, 1H), 7.32 (d, 1H,J = 2.1 Hz), 7.47 (d, 1H, J = 2.0 Hz), 8.54 (d, 1H, J = 2.1 Hz), 8.83 (d,1H, J = 2.2 Hz). LC-MS: Rt = 1.95 min; MS: m/z = 344 [M+1]+, 345[M+2]+.

4.1.8. 2-(3-Bromo-8-methylquinolin-6-yloxy)-N-tert-butyl-2-methylsulfanyl-acetamide (12)

Triethylamine (1.5 g, 15 mmol), 1-hydroxy-7-azabenzotriazole(2.0 g, 15 mmol), 1-ethyl-3-(3-dimethylamino-propyl)carbodiim-

3928 C. Lamberth et al. / Bioorg. Med. Chem. 22 (2014) 3922–3930

ide (2.8 g, 15 mmol) and 2-(3-bromo-8-methylquinoline-6-yloxy)-2-methylsulfanyl-acetic acid (11, 5.0 g, 15 mmol) were added con-secutively to a solution of tert-butylamine (1.1 g, 15 mmol) in70 ml of N,N-dimethylformamide. The reaction mixture was stirredfor 5 h at room temperature, then diluted with ethyl acetate andextracted with brine, saturated aqueous sodium bicarbonate solu-tion and water. The organic layer was dried over magnesium sul-fate and evaporated under reduced pressure, the residue waspurified by chromatography on silica gel, using ethyl acetate andhexane as eluents to deliver 2-(3-bromo-8-methylquinolin-6-yloxy)-N-tert-butyl-2-methylsulfanyl-acetamide (12, 3.9 g,9.8 mmol, 67%). 1H NMR (CDCl3): d = 1.43 (s, 9H), 2.19 (s, 3H),2.78 (s, 3H), 5.56 (s, 1H), 6.43 (br s, 1H), 7.00 (d, 1H, J = 2.1 Hz),7.31 (d, 1H, J = 2.1 Hz), 8.20 (d, 1H, J = 2.2 Hz), 8.82 (d, 1H,J = 2.0 Hz). LC-MS: Rt = 2.09 min; MS: m/z = 399 [M+1]+, 400[M+2]+.

4.1.9. 2-Methylsulfanyl-2-(8-methyl-3-vinylquinolin-6-yloxy)-acetic acid methyl ester (13)

Tetrakis(triphenylphosphine)palladium (130 mg, 0.11 mmol)was added to a solution of 2-(3-bromo-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (10, 2.0 g,5.6 mmol) and vinyltributyltin (1.8 g, 5.6 mmol) in 50 ml of tolu-ene at room temperature, The reaction mixture was heated to100 �C for 20 h, then cooled to room temperature. Saturated aque-ous sodium carbonate solution was added and the resulting mix-ture was stirred for 1 h at room temperature, subsequentlydiluted with ethyl acetate and then extracted with 5% aqueousammonium hydroxide solution and brine. The organic layer wasdried over magnesium sulfate and evaporated under reduced pres-sure, the residue was purified by chromatography on silica gel,using ethyl acetate and hexane as eluents to deliver 2-methylsulfa-nyl-(8-methyl-3-vinylquinolin-6-yloxy)acetic acid methyl ester(13, 0.8 g, 2.6 mmol, 47%). 1H NMR (CDCl3): d = 2.25 (s, 3H), 2.78(s, 3H), 3.89 (s, 3H), 5.47 (d, 1H, J = 11.2 Hz), 5.75 (s, 1H), 5.98 (d,1H, J = 17.8 Hz), 6.87 (dd, 1H, J = 11.1, 17.7 Hz), 7.06 (d, 1H,J = 2.1 Hz), 7.33 (d, 1H, J = 2.2 Hz), 7.97 (d, 1H, J = 2.0 Hz), 8.94 (d,1H, J = 2.1 Hz). LC-MS: Rt = 1.77 min; MS: m/z = 304 [M+1]+, 305[M+2]+.

4.1.10. N-(1-Cyano-2-methoxy-1-methyl-ethyl)-2-methylsulfanyl-2-(8-methyl-3-vinylquinolin-6-yloxy)-acetamide (14)

0.5 N sodium hydroxide (5 ml, 2.5 mmol) was slowly added at0 �C to a solution of 2-methylsulfanyl-(8-methyl-3-vinylquinolin-6-yloxy)acetic acid methyl ester (13, 0.6 g, 2.0 mmol) in 20 ml oftetrahydrofuran. The reaction mixture was stirred for 1 h at 0 �C,then 1 N hydrochloric acid was added to the remaining mixtureuntil pH 3 was reached. The mixture was extracted with ethyl ace-tate, the organic layer was washed with water and brine, dried overmagnesium sulfate and evaporated under reduced pressure to deli-ver 2-methylsulfanyl-(8-methyl-3-vinylquinolin-6-yloxy)aceticacid. This intermediate was dissolved in 17 ml of acetonitrile,and triethylamine (0.35 g, 3.4 mmol), 1-hydroxy-7-azabenzotria-zole (0.1 g, 0.8 mmol), O-(benzotriazolyl-1-yl)-N,N,N0,N0-tetram-ethyluronium tetrafluoroborate (0.24 g, 0.8 mmol) and 2-amino-3-methoxy-2-methyl-propionitrile (90 mg, 0.8 mmol) were addedconsecutively. The reaction mixture was stirred for 16 h at roomtemperature, then diluted with ethyl acetate and extracted withbrine, saturated aqueous sodium bicarbonate solution and water.The organic layer was dried over magnesium sulfate and evapo-rated under reduced pressure. The residue was purified by chroma-tography on silica gel, using ethyl acetate and cyclohexane aseluents to deliver N-(1-cyano-2-methoxy-1-methylethyl)-2-meth-ylsulfanyl-2-(8-methyl-3-vinylquinolin-6-yloxy)-acetamide (14,

0.38 g, 1.0 mmol, 49% over two steps). 1H NMR (CDCl3): d = 1.82(d, 3H), 2.23 (s, 3H), 2.81 (s, 3H), 3.51 (d, 3H), 3.68–3.82 (m, 2H),5.49 (d, 1H, J = 11.1 Hz), 5.74 (d, 1H), 5.99 (d, 1H, J = 17.7 Hz),6.88 (dd, 1H, J = 11.0, 17.7 Hz), 7.12 (d, 1H, J = 2.0 Hz), 7.28 (d,1H, J = 2.1 Hz), 8.00 (d, 1H, J = 2.0 Hz), 8.97 (d, 1H, J = 2.2 Hz). LC-MS: Rt = 1.71 min; MS: m/z = 386 [M+1]+, 387 [M+2]+.

4.1.11. 2-(3-Ethynyl-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (15)

Trimethylsilylacetylene (4.1 g, 42 mmol) was added dropwiseto a degassed solution of 2-(3-bromo-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (10, 10 g, 28 mmol),copper iodide (0.2 g, 1.4 mmol), diisopropylamine (5.7 g, 56 mmol)and bis(triphenylphosphine)palladium(II) dichloride (1.0 g,1.4 mmol) in 150 ml of tetrahydrofuran. The reaction mixturewas stirred for 24 h at room temperature and then washed withbrine. The aqueous phase was extracted with ethyl acetate, thecombined organic layer washed with water, dried over magnesiumsulfate and evaporated under reduced pressure. The residue waspurified by chromatography on silica gel, using ethyl acetate andheptane as eluents to deliver 2-methylsulfanyl-(8-methyl-3-trim-ethylsilanylethynyl-quinolin-6-yloxy)-acetic acid methyl ester.This intermediate was dissolved in 200 ml of methanol and treatedwith potassium carbonate (8.1 g, 59 mmol). The reaction mixturewas stirred for 30 min at room temperature, then poured on satu-rated aqueous sodium bicarbonate solution and extracted withethyl acetate. The organic layer was dried over magnesium sulfateand evaporated under reduced pressure, the residue was purifiedby chromatography on silica gel, using ethyl acetate and heptaneas eluents to deliver 2-(3-ethynyl-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (15, 4.6 g, 15 mmol, 54%)1H NMR (CDCl3): d = 2.24 (s, 3H), 2.78 (s, 3H), 3.27 (s, 1H), 3.88(s, 3H), 5.73 (s, 1H), 7.02 (d, 1H, J = 2.1 Hz), 7.37 (d, 1H,J = 2.1 Hz), 8.16 (d, 1H, J = 2.1 Hz), 8.85 (d, 1H, J = 2.0 Hz). LC-MS:Rt = 1.87 min; MS: m/z = 302 [M+1]+, 303 [M+2]+.

4.1.12. 2-(3-Ethynyl-8-methylquinolin-6-yloxy)-N-(2-methoxyimino-1,1-dimethyl-ethyl)-2-methyl-sulfanyl-acetamide (2)

2 N sodium hydroxide (3.1 ml, 6.3 mmol) was added to a solutionof 2-(3-ethynyl-8-methylquinolin-6-yloxy)-2-methylsulfanyl-ace-tic acid methyl ester (15, 1.2 g, 4.2 mmol) in 50 ml of ethanol. Thereaction mixture was stirred for 1 h at room temperature, then 2 Nhydrochloric acid was added to the remaining mixture until pH 3was reached. The mixture was poured on brine and extracted withethyl acetate. The organic layer was dried over magnesium sulfateand evaporated under reduced pressure to deliver 2-(3-ethynyl-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid. This inter-mediate was dissolved in 12 ml of N,N-dimethylformamide and,triethylamine (0.4 g, 4.4 mmol), 1-hydroxy-7-azabenzotriazole (0.6 g,4.4 mmol), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (0.8 g,4.4 mmol) and 2-amino-2-methyl-propionaldehyde O-methyl-oximehydrochloride(0.7 g,4.4 mmol)wereaddedconsecutively.Thereactionmixture was stirred for 16 h at room temperature, then diluted withethyl acetate and extracted with brine. The organic layer was dried overmagnesium sulfateand evaporated under reduced pressure, the residuewas purified by chromatography on silica gel, using ethyl acetate andhexane as eluents to deliver 2-(3-ethynyl-8-methylquinolin-6-yloxy)-N-(2-methoxyimino-1,1-dimethyl-ethyl)-2-methylsulfanyl-acet-amide (2, 0.9 g, 2.3 mmol, 56% over two steps). 1H NMR (CDCl3):d = 1.59 (s, 3H), 1.62 (s, 3H), 2.21 (s, 3H), 2.79 (s, 3H), 3.31 (s, 1H),3.89 (s, 3H), 5.66 (s, 1H), 7.06 (d, 1H, J = 2.1 Hz), 7.33 (d, 1H,J = 2.2 Hz), 7.59 (s, 1H), 8.17 (d, 1H, J = 2.2 Hz), 8.85 (d, 1H,J = 2.1 Hz). LC-MS: Rt = 1.88 min; MS: m/z = 386 [M+1]+, 387[M+2]+.

C. Lamberth et al. / Bioorg. Med. Chem. 22 (2014) 3922–3930 3929

4.1.13. 2-(3,8-Dimethylquinolin-6-yloxy)-2-methyl-sulfanyl-acetic acid methyl ester (16)

A degassed solution of 2-(3-bromo-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (10, 4.0 g, 11 mmol),trimethylboroxine (1.5 g, 12 mmol), potassium carbonate (4.7 g,34 mmol) and tetrakis(triphenylphosphine)palladium(0) (1.3 g,1.1 mmol) in 90 ml of dioxane was heated to 100 �C for 5 h. Thereaction mixture was cooled to room temperature, then dilutedwith ethyl acetate and extracted with water. The organic layerwas washed with brine, dried over magnesium sulfate and concen-trated under reduced pressure. The residue was purified by chro-matography on silica gel, using ethyl acetate and cyclohexane aseluents to deliver 2-(3,8-dimethylquinolin-6-yloxy)-2-meth-ylsulfanyl-acetic acid methyl ester (16, 1.7 g, 5.7 mmol, 51%). 1HNMR (CDCl3): d = 2.26 (s, 3H), 2.51 (s, 3H), 2.78 (s, 3H), 3.87 (s,3H), 5.74 (s, 1H), 6.99 (d, 1H, J = 2.1 Hz), 7.31 (d, 1H, J = 2.1 Hz),7.82 (d, 1H, J = 2.2 Hz), 8.69 (d, 1H, J = 2.1 Hz). LC-MS: Rt = 1.45 -min; MS: m/z = 292 [M+1]+, 293 [M+2]+.

4.1.14. 2-(3,8-Dimethylquinolin-6-yloxy)-N-(4-methoxy-1,1-dimethyl-but-2-ynyl)-2-methylsulfanyl-acetamide (17)

1 N sodium hydroxide (7.5 ml, 7.5 mmol) was added to a solu-tion of 2-(3,8-dimethylquinolin-6-yloxy)-2-methylsulfanyl-aceticacid methyl ester (16, 1.7 g, 5.7 mmol) in 25 ml of tetrahydrofuranat 0 �C. The reaction mixture was stirred for 1 h at room tempera-ture, then 2 N hydrochloric acid was added to the remaining mix-ture until pH 3 was reached. The mixture was poured on brine andextracted with ethyl acetate. The organic layer was dried over mag-nesium sulfate and evaporated under reduced pressure to deliver2-(3,8-dimethylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid.This intermediate was dissolved in 80 ml of acetonitrile and trieth-ylamine (2.0 g, 20 mmol), 1-hydroxy-7-azabenzotriazole (1.1 g,8.5 mmol), O-(benzotriazolyl-1-yl)-N,N,N0,N0-tetramethyluroniumtetrafluoroborate (2.7 g, 8.5 mmol) and 4-methoxy-1,1-dimethyl-but-2-ynylamine hydrochloride (1.4 g, 8.5 mmol) were added con-secutively. The reaction mixture was stirred for 16 h at room tem-perature, then diluted with ethyl acetate and extracted with brine,saturated aqueous sodium bicarbonate solution and water. Theorganic layer was dried over magnesium sulfate and evaporatedunder reduced pressure. The residue was purified by chromatogra-phy on silica gel, using ethyl acetate and cyclohexane as eluents todeliver 2-(3,8-dimethylquinolin-6-yloxy)-N-(4-methoxy-1,1-dimethyl-but-2-ynyl)-2-methylsulfanyl-acetamide (17, 1.7 g, 4.6 mmol, 82% over two steps). 1H NMR (CDCl3): d = 1.74 (s, 6H), 2.21(s, 3H), 2.50 (s, 3H), 2.78 (s, 3H), 3.38 (s, 3H), 4.13 (s, 2H), 5.63(s, 1H), 6.78 (br s, 1H), 7.03 (d, 1H, J = 2.0 Hz), 7.24 (d, 1H,J = 2.1 Hz), 7.82 (d, 1H, J = 2.0 Hz), 8.70 (d, 1H, J = 2.2 Hz). LC-MS:Rt = 1.54 min; MS: m/z = 387 [M+1]+, 388 [M+2]+.

4.1.15. 2-(3-Cyclopropyl-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (18)

Tetrakis(triphenylphosphine)palladium(0) (0.3 g, 0.3 mmol)was added to a degassed solution of 2-(3-bromo-8-methylquino-lin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (10, 2.0 g,5.6 mmol), cyclopropyl boronic acid (0.6 g, 7.3 mmol) and potas-sium phosphate tribasic (4.1 g, 19 mmol) in 40 ml of toluene and2 ml of water. The reaction mixture was heated to 100 �C for 5 h,then cooled to room temperature, diluted with ethyl acetate andextracted with saturated aqueous sodium bicarbonate solution.The organic layer was washed with brine, dried over magnesiumsulfate and evaporated under reduced pressure. The residue waspurified by chromatography on silica gel, using ethyl acetate andcyclohexane as eluents to deliver 2-(3-cyclopropyl-8-methylquin-olin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (18,0.45 g, 1.4 mmol, 25%). 1H NMR (CDCl3): d = 0.87 (q, 2H), 1.12 (q,2H), 2.04–2.11 (m, 1H), 2.27 (s, 3H), 2.78 (s, 3H), 3.89 (s, 3H),

5.73 (s, 1H), 7.01 (d, 1H, J = 2.2 Hz), 7.30 (d, 1H, J = 2.1 Hz), 7.63(d, 1H, J = 2.0 Hz), 8.69 (d, 1H, J = 2.1 Hz). LC-MS: Rt = 1.70 min;MS: m/z = 318 [M+1]+, 319 [M+2]+.

4.1.16. N-(2-Chloroallyl)-2-(3-cyclopropyl-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetamide (19)

0.5 N sodium hydroxide (2.6 ml, 1.3 mmol) was added to a solu-tion of 2-(3-cyclopropyl-8-methylquinolin-6-yloxy)-2-methylsulfanyl-acetic acid methyl ester (18, 0.32 g, 1.0 mmol) in 10 ml of tetra-hydrofuran at 0 �C. The reaction mixture was stirred for 2 h at roomtemperature, then 2 N hydrochloric acid was added to the remainingmixture until pH 3 was reached. The mixture was poured on brineand extracted with ethyl acetate. The organic layer was dried overmagnesium sulfate and evaporated under reduced pressure todeliver 2-(3-cyclopropyl-8-methylquinolin-6-yloxy)-2-meth-ylsulfanyl-acetic acid. This intermediate was dissolved in 5 ml ofN,N-dimethylformamide and, triethylamine (0.25 g, 2.5 mmol),1-hydroxy-7-azabenzotriazole (0.17 g, 1.2 mmol), 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (0.24 g, 1.2 mmol) and 2-chlo-roallylamine (0.11 g, 1.2 mmol) were added consecutively. The reac-tion mixture was stirred for 16 h at room temperature, then dilutedwith ethyl acetate and extracted with saturated aqueous sodiumbicarbonate solution. The organic layer was washed with brine,dried over magnesium sulfate and evaporated under reduced pres-sure, the residue was purified by chromatography on silica gel, usingethyl acetate and cyclohexane as eluents to deliver N-(2-chloroal-lyl)-2–3-cyclopropyl-8-methylquinolin-6-yloxy)-2-methylsulfa-nyl-acetamide (19, 0.21 g, 0.55 mmol, 56% over two steps). 1H NMR(CDCl3): d = 0.73–0.81 (m, 2H), 1.01–1.08 (q, 2H), 1.96–2.02 (m, 1H),2.14 (s, 3H), 2.69 (s, 3H), 4.03 (dd, 1H, J = 11.2, 17.3 Hz), 4.20 (dd, 1H,J = 11.0, 17.3 Hz), 5.27 (s, 1H), 5.36 (s, 1H), 5.65 (s, 1H), 6.92 (br s, 1H),6.97 (d, 1H, J = 2.2 Hz), 7.18 (d, 1H, J = 2.1 Hz), 7.55 (d, 1H, J = 2.0 Hz),8.61 (d, 1H, J = 2.1 Hz). LC-MS: Rt = 0.87 min; MS: m/z = 377 [M+1]+,379 [M+3]+.

4.2. Biology

4.2.1. Phytophthora infestans/tomato (action against late blighton tomato)

Three-week old tomato plants cv. Roter Gnom were sprayed in aspray chamber with the formulated test compound diluted inwater. The test plants were inoculated by spraying them with asporangia suspension two days after application. The inoculatedtest plants were incubated at 18 �C with 14 h light/day and 100%rh in a growth chamber and the percentage leaf area covered bydisease was assessed when an appropriate level of diseaseappeared on untreated check plants (5–7 days after application).

4.2.2. Mycosphaerella graminicola (Septoria tritici)/wheat(action against leaf blotch on wheat)

Two-week old wheat plants cv. Riband were sprayed in a spraychamber with the formulated test compound diluted in water. Thetest plants were inoculated by spraying a spore suspension onthem one day after application. After an incubation period of1 day at 22 �C/21 �C (day/night) and 95% rh, the inoculated testplants were kept at 22 �C/21 �C (day/night) and 70% rh in a green-house. Efficacy was assessed directly when an appropriate level ofdisease appeared on untreated check plants (16–19 days afterapplication).

4.2.3. Uncinula necator (Erysiphe necator)/grape (action againstpowdery mildew on grape)

Five-week old grape seedlings cv. Gutedel were sprayed in aspray chamber with the formulated test compound diluted inwater. The test plants were inoculated by shaking plants infectedwith grape powdery mildew above them 1 day after application.

3930 C. Lamberth et al. / Bioorg. Med. Chem. 22 (2014) 3922–3930

The inoculated test plants were incubated at 24/22 �C (day/night)and 70% rh under a light regime of 14/10 h (light/dark) and thepercentage leaf area covered by disease was assessed when anappropriate level of disease appeared on untreated check plants(7–9 days after application).

Acknowledgments

The authors gratefully acknowledge the excellent syntheticcontribution of Myriam Baalouch, Armando Cicchetti, AntoniettaCrisante, Thomas Fischer, Florian Garo, Thomas Grether, NikolaKokosar, Jerome Sachet, Grit Schade and Thomas Steffen.

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