Synthesis and anti-HIV-1 evaluation of new Sonogashira-modified emivirine (MKC-442) analogues

Article abstract of DOI:10.1002/hlca.200900039

The MKC-442 analogue 6-(3,5-dimethylbenzyl)-5-ethyluracil substituted with a (propargyloxo)-methyl group at N(1) has previously been found highly active against HIV-1. The C≡C bond in the substituent at N(1) is here utilized in a series of chemical reacti

Full text of DOI:10.1002/hlca.200900039

Helvetica Chimica Acta – Vol. 92 (2009)
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Synthesis and Anti-HIV-1 Evaluation of New Sonogashira-Modified
Emivirine (MKC-442) Analogues
by Krzysztof Danel1)a), Per T. Jørgensena), Erik B. Pedersen*a), Paolo La Collab), Gabriella Collub),
and Roberta Loddob)
a) Nucleic Acid Center2), Department of Physics and Chemistry, University of Southern Denmark,
Campusvej 55, DK-5230 Odense M (phone: þ 4565502555; fax: þ 4566158780; e-mail: ebp@ifk.sdu.dk)
b) Dipartimento di Scienze e Tecnologie Biomediche, Sezione di Microbiologia e Virologia Generale e
´
Biotecnologie Microbiche, Universita di Cagliari, Cittadella Universitaria, I-09042 Monserrato
The MKC-442 analogue 6-(3,5-dimethylbenzyl)-5-ethyluracil substituted with a (propargyloxo)-
methyl group at N(1) has previously been found highly active against HIV-1. The CꢀC bond in the
substituent at N(1) is here utilized in a series of chemical reactions in order to develop new agents with
higher activity against HIV-1-resistant mutants. The syntheses involved Pd-catalyzed C,C-coupling
reactions, addition of disulfides, and click chemistry on the terminal CꢀC bond as well as addition of
bromine to the so formed internal CꢀC bonds. Sonogashira coupling were performed with silyl-
derivatized iodobenzyl alcohols which, after deprotection, were oxidized to aldehydes by means of IBX.
The isomeric alcohol 37 was obtained in the Sonogashira reaction of propargyl alcohol with the N(1)-
substituted (4-iodobenzyloxy)methyl derivative of the above mentioned uracil. Compound 37 turned out
to be the most effective compound against problematic HIV-1 mutants. The general observation in the
present work is that a combination of alkyne and aryl in the substituent at N(1) leads to highly active
compounds against HIV-1.
Introduction. – About 25 years have now elapsed since the discovery that the
causative agent of acquired immunodeficiency syndrome (AIDS), pandemic disease
affecting millions of people all over the world, is HIV-1 virus [1][2]. The life cycle of
HIV-1 virus has been recognized in details, and nearly all events in its replicative cycle
can be effectively targeted [3][4]. One of the most important step during the
replicative process of the virus is the reverse transcription mediated by the virally
encoded reverse transcriptase (RT) enzyme. Two different classes of compounds
possess powerful ability to terminate this important process, nucleoside RT inhibitors
ꢀnukesꢁ (NRTIs) and non-nucleoside RT inhibitors (NNRTIs). The former can inhibit
the enzyme competitively, as chain terminators; the latter are allosteric inhibitors
interfering by a noncompetitive mechanism, binding to a pocket near the polymerase
active site [5 – 7]. The NNRTIs are less toxic than NRTIs; however, their prolonged use
lead to rapid development of drug-resistant viral strains. The clinically most important
mutations of HIV-1 RToccur in amino acids Lys-103, Tyr-181, and Tyr-188. The K103N
1
)
On leave from Department of Chemistry, Agricultural University, Balicka St. 122, PL-30-149
Krakow.
2
)
A research center founded by The Danish National Research Foundation for studies on nucleic acid
chemical biology.
ꢂ 2009 Verlag Helvetica Chimica Acta AG, Zꢃrich

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mutation is very destructive to efficacy of most NNRTIs. In wild-type HIV-1 virus, the
entrance to the pocket is ꢀopenedꢁ, whereas, in the mutated K103N type, it is ꢀclosedꢁ,
because of additional H-bonding between the phenol of Tyr-188 and the amine of 103
asparagine [8]. The Y181C mutation is responsible for the lost of binding energy of
NNRTIs with aromatic plane of Tyr-181 [9]. In 2002, one of us synthesized a number of
extremely potent NNRTIs of the MKC-442 type (Fig.) against wild-type HIV-1 virus
with unprecedented selective index (SI) of 5,000,000 [10]. Moreover, their potency
against the clinically important Y181C and K103N mutant strains was also enhanced.
Among the huge number of newly synthesized anti-HIV compounds, 1a (cf. Scheme 1)
[10] is an example of an anti-HIV compounds containing a terminal CꢀC bond, the
first one being the NRTI 4’-ethynylthymidine, which later has been followed by 4’-E-dC
(4’-Ethynyl-2’-deoxycytidine; Fig.) [11].
Figure. Structures of MKC-442, TNK-651, and 4’-Ethynyl-2’-deoxycytidine
As there seems to be structural freedom of the substituents at N(1) in MKC-442-
type drugs, it was tempting to perform some chemistry on the terminal CꢀC bond of
1a. The obvious choice seems to be Sonogashira reaction, which has been gaining
tremendous importance during the last decade [12][13]. Some recent examples showed
coupling at the C(5) of the pyrimidine ring with different alkynes [14 – 18]. However,
we decided to exploit the potential of palladacycles to modify the terminal CꢀC bond
of 1a, because this is one of the only sites where new important modifications may be
performed within the framework of the uracil ring [19][20]. One of our first attempts to
use Pd-promoted cross-coupling reactions to generate new HIV-1 inhibitors from 1a
has been successfully demonstrated in our recent work [21]. Therefore, the current
study is the continuation of our previous efforts to synthesize novel MKC-442
analogues with improved activity against resistant mutants.
Results and Discussion. – Chemistry. The lead compounds 1a and 1b (Scheme 1) to
be submitted to Sonogashira reactions were synthesized according to the procedure
reported in [10]. When performing Sonogashira reaction, a problem of homocoupling
reaction aroused [13]. We applied a general procedure for the cross-coupling reaction,
namely, dry Et3N, [PdCl2(PPh3)2], CuI, and aryl iodide (ArI) at room temperature
under an Ar atmosphere. In most cases, we obtained the expected cross-coupled
products in high yield and did not isolate the undesired homocoupled alkynes [21].
However, when unprotected phenols, benzyl alcohols, and anilines were used, the yield
of cross-coupled products decreased significantly. It was especially difficult to prepare

Helvetica Chimica Acta – Vol. 92 (2009)
1387
Scheme 1
i
a) [PdCl2(PPh3)2], CuI, ArI, Et3N, r.t. b) SnCl2 · H2O, AcOEt, reflux. c) KOH, PrOH, reflux. d)
(BrCH2CO)2O, THF, r.t. e) [PdCl2(PPh3)2], CuI, Et3N, r.t. f) TBAF, THF. g) 2-Iodoxybenzoic acid
(IBX), AcOEt, r.t.

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Helvetica Chimica Acta – Vol. 92 (2009)
the interesting 4-aminophenyl derivative 6a directly from unprotected 4-iodoaniline
and alkyne 1a.
Related coupling reactions have been described in the literature with a reasonable
yield of ca. 60% by other groups [22]. However, in our attempt to synthesize the 4-
aminophenyl derivatives 6a and 6b, the yield for the coupled product did not exceed
20%, and it was accompanied mostly by homocoupled counterpart. Thus, to prepare 6a
we reacted the electron-deficient 1-iodo-4-nitrobenzene with 1a under standard
Sonogashira condition, and we obtained the corresponding nitrobenzene ethyne 9a in a
high yield (Scheme 1). The reduction of the NO2 moiety on 9a was accomplished by
treatment with SnCl2 according to the procedure reported in [23]. However, instead of
the expected aminophenyl compound 6a, 5-ethyl-6-(3,5-dimethylbenzyl)uracil [24] was
formed as the only product (Scheme 1). A related N-dealkylation reaction has recently
been developed by Chen et al. [25] by using a mixture of hypophosphorous acid and
iodine in AcOH. However, we solved these difficulties by protecting the 4-iodoaniline
with (CF3CO)2O [26][27] and its trifluoroacetamide derivative was coupled with 1a
and 1b to give the amides 16a and 16b, respectively, in high yields. The free amines 6a
and 6b were obtained after refluxing the amides with KOH/i-PrOH in 82 – 87% isolated
yield (Scheme 1). We have previously attempted to synthesize potential irreversible
inhibitors of HIV-1 in our laboratory [28 – 31]. Since bromoacetamides as nonsteroidal
anti-inflammatory drugs (NSAID) analogues were employed successfully to map the
active sites of hydroxysteroid dehydrogenases [32][33], we saw a potential in this
functionality. Encouraged by a recent report on benzyl halides Pd-coupling with
terminal ethynes [34], we used the method to couple bromoacetylated 4-iodoaniline
directly to compound 1a (Scheme 1). Unfortunately, the yield (<2%) of the main
product 17a was not satisfactory after tedious workup. The 4-aminophenyl compounds
6a and 6b were, therefore, converted to the corresponding bromoacetamides 17a and
17b, respectively, by means of (BrCH2CO)2O in THF to deliver the potential
irreversible inhibitors of HIV-1 RT in reasonable yield (Scheme 1).
As mentioned above, the electron-rich aryl iodophenols and iodobenzyl alcohols
had to be conveniently protected with the tBuPh2Si (TBDPS) groups (see Exper. Part)
prior to coupling with 1a to avoid homocoupled products [13]. We also synthesized the
silyl-protected iodopropargyl alcohol [35] to extend acetylenic fragment of 1a
producing 18a. Deprotection of the resulting silyl derivatives was accomplished
quantitatively by means of Bu4NF (TBAF) in THF to give 23a – 27a (Scheme 1).
Next, we performed some modifications of the CꢀC bonds in 1a and 1b
(Scheme 2). The importance of vinylic chalcogenides has been recognized recently
[36][37]. Addition of Ph2S2 to 1a according to a published protocol resulted in the (Z)-
isomer 30 [37]. Click chemistry is another powerful synthetic tool with an increasing
popularity [38 – 41]. Recently, we took advantage of Huisgen 1,3-dipolar cycloaddition
of azides to CꢀC bond to synthesize a double drug in reaction of 1a and AZT [21].
Here, we prepared a series of triazolyl derivatives 31 – 34 (Scheme 2). Only 1,4-
regioisomeric products were formed, which were established by comparing their NMR
spectral data with those of the related compounds already published [42][43].
Electrophilic addition of 1 equiv. of Br2 to the CꢀC bond in 2a and 2b at 08 in dry
CH2Cl2 gave nearly quantitatively one isomer, assumed to be the (E)-isomers of 35a
and 35b, respectively. This is in agreement with other reports [44][45].

Helvetica Chimica Acta – Vol. 92 (2009)
1389
Scheme 2
a) Ph2S2, Pd(PPh3)4, PPh3. b) Click chemistry. c) Br2, CH2Cl2, 08.
A powerful TNK-651 (Fig.) analogue, 36, obtained in our laboratory recently
[21][46], was treated with propargyl alcohol (¼ prop-2-yn-1-ol) to deliver compound
37 as the isomeric analogue of 26a and 27a (Scheme 3). The alcohols 26a, 27a, and 37
Scheme 3
a) Propargyl alcohol, [PdCl2(PPh3)2], CuI, Et3N, r.t. b) IBX, AcOEt, r.t.

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Helvetica Chimica Acta – Vol. 92 (2009)
were subsequently oxidized to the corresponding aldehydes 28a, 29a, and 38,
respectively, with 2-iodoxybenzoic acid (IBX) in AcOEt (Schemes 1 and 3) [47][48].
Biological Results and Discussion. – The HIV-1 strain HTLV-IIIB in MT-4 cells
were used in our assay to investigate the anti-HIV-1 activity and cytotoxicity of the
MKC-442 analogues synthesized in the present study. The results are summarized in the
Table together with those of MKC-442 and efavirenz. The activity data of the two lead
ethynyl compounds 1a and 1b are also included for comparison. The first series of
alkynes concerns with substituted MKC-442 analogues having big aryl rings attached to
the CꢀC bond of 1a and 1b, i.e., phenanthren-1-yl, naphthalen-1-yl, and 9H-fluoren-2-
yl, 3a, 4b, and 5b, respectively, and electron-withdrawing or -donating substituents at
the phenyl ring, compounds 6a – 12a. Additionally, compounds 13a – 15a are charac-
terized by the presence of heterocyclic moieties (thiophene, pyridine, and pyrazine) as
the extension of the terminal CꢀC bond. The rationale to prepare 1b lies in the fact that
one of the most anti-HIV-1-active diarylpyrimidine (DAPY) analogues bears a 2,4,6-
trimethyl moiety attached to the pyrimidine ring [49]. Furthermore, within the HEPT
(1-[(2-hydroxyethoxy)methyl]-6-(phenylsulfanyl)thymine) series, there are reports
related to the modification at C(2) of the phenylsulfanyl ring [50 – 52]. The biological
results presented here reveal clearly that this approach did not enhance the antiviral
activity. However, when we compare the lead compound 1b (EC50 ¼ 0.4 mm) with the
corresponding 4-aminophenyl analogue 6b (EC50 ¼ 0.07 mm), we find nearly a six times
higher activity of the latter. Interestingly, when compound 6b is converted to the
trifluoro- or bromoacetamido derivatives 16b, 17b, respectively, the potency of both
compounds (EC50 ¼ 0.09 and 0.07 mm, resp.) is preserved. The same tendency is also
observed in the series 6a, 16a, and 17a (EC50 ¼ 0.007, 0.007, and 0.008 mm, resp.). It
seems that the amides are easily hydrolyzed in the biological assay to regenerate the
active compounds 6a and 6b. In this way, one can consider these derivatives as prodrugs
of 6a and 6b. Similar prodrugs against solid tumors have been synthesized recently
[53]. From the activity data of 17a and 17b, it is not possible to conclude that the
bromoacetamido group is a target for nucleophilic attack to yield carboxymethylated
RT. The data collected in the Table clearly confirm the advantages of the 3,5-dimethyl
substitution pattern at the 6-benzyl ring [20][50] which resulted in the following ratios:
EC50 (2a)/EC50 (2b) ¼ 100; EC50 (16a)/EC50 (16b) ¼ 13; EC50 (6a)/EC50 (6b) ¼ 10; EC50
(17a)/EC50 (17b) ¼ 9, for instance. It seems that an amine moiety compensate to some
extent for the loss of activity of 2-Me substitution at the 6-benzyl ring. The most active
modifications of 1a within the series 2a – 15a are: 2a (EC50 ¼ 0.004 mm), 6a (EC50 ¼
0.007 mm), and 13a (EC50 ¼ 0.008 mm). They are also active against mutated forms of
HIV-1. Compound 11a, bearing a 3-cyanophenyl ring, though slightly less potent than
the previous analogues, is also active against mutants. Recently published SJ-3366
analogues, containing a [(phenylpropynyl)oxy]methyl chain at N(1) of the uracil ring
are less potent than 2a [54]. Compound 3a, an inhibitor with a phenanthren-9-yl
attached ring, is 100 times less potent than the parent compound 1a, and it is not active
against mutants at subtoxic concentrations. The next group of compounds comprises
silyl ethers 18a – 22a, phenols 24a and 25a, primary alcohols 23a, 26a, 27a, and 37, and
the corresponding aldehydes 28a, 29a, and 38. The least active compounds within this
group of inhibitors are the silyl ethers. On the other hand, they are less cytotoxic (CC50

Helvetica Chimica Acta – Vol. 92 (2009)
1391
Table. Cytotoxicity and Anti-HIV-1 Activity in MT-4 Cells
Compound
CC50 [mm]a)
SIb)
EC50 [mm]c)
Wild type
EFVR
Y181C
K103N þ Y181C
1a
1b
2a
2b
3a
4b
5b
6a
6b
> 100
33
> 33333
82
0.003
0.4
0.004
0.4
0.3
1.8
16
> 33
12
> 26
> 80
> 34
> 50
6
> 39
100
> 10
> 7
0.2
> 33
2
1.2
> 33
6
> 26
> 80
> 34
> 50
2.4
> 39
18
> 10
> 7
15
8.5
> 70
5
> 16
11
1.6
> 45
2.3
> 41
> 76
> 100
6
> 100
> 100
1.7
6
5
2
1.8
4
6.4
12
100
21
> 48
14
11
> 26
0.9
0.6
8
> 100
0.3
30
26
80
34
50
40
39
85
10
7
25
39
70
38
16
30
7500
65
267
19
4
5714
557
4250
167
233
417
> 26
> 80
> 34
> 50
1.8
> 39
100
> 10
> 7
> 25
5
12
0.007
0.07
0.02
0.06
0.03
0.06
0.01
0.8
0.008
0.01
0.02
0.007
0.09
0.008
0.07
2
7a
8a
9a
10a
11a
12a
13a
14a
15a
16a
16b
17a
17b
18a
19a
20a
21a
22a
23a
24a
25a
26a
27a
28a
29a
30
> 25
3900
88
7.5
> 70
15
16
14
9
> 45
7
> 41
> 76
> 100
60
4750
1600
1500
5143
500
4875
586
38
2.5
> 16
11
1.5
> 45
1.3
13
> 76
20
7.6
20
23
0.8
3
4
1
36
45
39
41
76
> 100
> 100
> 100
> 100
40
> 59
> 200
> 71
> 167
5000
2500
2200
6000
11333
4500
2100
1000
> 12500
> 10000
1600
771
1.7
0.5
1.4
0.6
5.4
> 100
> 100
17
0.008
0.008
0.02
0.007
0.003
0.01
0.02
0.06
0.008
0.01
0.03
0.07
0.07
0.6
20
44
42
34
45
42
60
20
11
10
7
14
13
> 60
62
6
4
4
12
1.6
1.7
3.6
7
31
32
33
34
35a
35b
36
37
38
> 100
> 100
48
20
2.6
2.8
3.2
1.4
8
0.1
0.07
1
54
36
26
19
41
44
514
43
> 26
2714
6833
550
> 3333
15000
0.007
0.006
0.08
0.03
0.002
0.8
0.5
5.5
100
3
MKC-442
EFV
> 100
30
20
0.008
a) Compound dose required to achieve 50% protection of MT-4 cells from HIV-1-induced cytopatho-
genicity, as determined by the MTT method. The symbol (>) indicates that the CC50 value was not
reached at the highest concentration tested. b) Selectivity index: ratio CC50/EC50. CC50 and EC50 are
expressed as the mean values of at least three separate experiments. Variation among triplicate samples
c
was less than 10%. ) Compound dose required to reduce the viability of mock-infected cells by 50%, as
determined by the MTT method.

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Helvetica Chimica Acta – Vol. 92 (2009)
values > 100 mm). It is easily to notice that 3-substituted analogues are more potent
than the corresponding 4-substituted analogues. Interestingly, compound 20a shows
almost the same activity against all tested strains. Also introducing acetylene linkage
instead of benzene ring did not influence the activity of the silyl ether 18a.
Deprotection of the silyl ethers resulted in very active antiviral compounds 23a – 27a.
Their activity is of the same order as the aniline 6a, as well as its acetamides 16a and
17a. All alcohols are also active against the mutants Y181C and K103N þ Y181C. In our
group, we have recently synthesized the TNK-651 analogue 36 [21] which proved to be
extremely potent against HIV-1. Therefore, its propargylation was straightforward, and
this led to a powerful inhibitor 37 with an excellent profile against HIV-1 and its
mutants. It is isomeric with the two alcohols 26a and 27a. Oxidation of alcohols gave
compounds 28a, 29a, and 38 with a CHO group which may react chemically with OH or
amino groups within the non-nucleoside binding site (NNBS). However, their activity
was of the same order as their corresponding alcohols, and, therefore, no additional
binding could be postulated.
The pharmacological activities of the bis(phenylsulfanyl) derivative 30, and the
dibromo analogues 35a and 35b are not superior compared to the lead 1a; however,
their potency is of the same magnitude as the lead compound MKC-442, except of 35b,
which is 20 times less potent than MKC-442. The pharmacological profile of the
triazoles 31 – 34 is remarkable. Although with comparable activity, the comparison
between 14a and 31 reveals less toxicity of the latter. When we introduced a CH2 linker
into the triazole ring, we obtained a better profile against mutant RTs than 14a and 31.
Conclusions. – In summary, we synthesized a series of novel NNRTIs inhibitors with
improved activity against resistant mutants. They all were derived from the lead
compound 1a by Sonogashira reaction and click chemistry. Their activity was in most
cases better than MKC-442. Our study confirmed that there are many possibilities for
changing the substituent at N(1) of MKC-442, which improves the chances of finding
new compounds with favorable activity/toxicity profiles. As continuation of our
previous work [10], we obtained a powerful inhibitor 37 with promising activity against
HIV-1 and its mutants. The presence of H-bond donating groups (OH, CH2OH, and
NH2) as well as H-bond acceptors (triazole, pyridine, CHO, and acetamides) is
beneficial for enhancing the interaction between inhibitors and the RT at the NNBS.
This work received funding from the European Communityꢀs Sixth Framework Programme under
contract number LSHP-CT-2004-503162 (Selection and development of microbicides for mucosal use to
prevent sexual HIV transmission/acquisition)3)
Experimental Part
General. Reactions were in general carried out under Ar. MeCN was dried over 3-ꢄ molecular
sieves. Compound 1a was prepared as described by one of us [10]. tBuPh2Si (TBDPS) groups in place of
tBuMe2Si (TBDMS) groups were used to protect propargyl alcohol [35], aryl-iodophenols, and
3
)
The information in this document is provided as it is, and no guarantee or warranty is given that the
information is fit for any particular purpose. The user thereof uses the information as its sole risk
and liability.

Helvetica Chimica Acta – Vol. 92 (2009)
1393
iodobenzyl alcohols [55][56]. 2-Iodo-9H-fluorene [57] and 2-bromo-N-(4-iodophenyl)acetamide [58]
were synthesized according to the published protocols. Silica gel (SiO2; 0.040 – 0.063 mm) used for
column chromatography (CC) and anal. SiO2 TLC plates 60 F254 were purchased from Merck. Solvents
for chromatography were bought as HPLC grade or distilled prior to use. M.p.: Bꢁchi melting-point
apparatus. NMR Spectra: Varian Gemini 2000 spectrometer at 300 and 75 MHz for 1H and 13C, resp., with
Me4Si as an internal standard. MALDI-MS: 4.7 Tesla HiRes MALDI Ultima (IonSpec, Irvine, CA)
Fourier Transform Ion Cyclotron Resonance (FTICR) mass spectrometer. Elemental analyses: at H. C.
Ørsted Institute, University of Copenhagen; analyses were within ꢁ 0.4% of the theoretical values.
5-Ethyl-6-(2-methylbenzyl)-1-[(prop-2-ynyloxy)methyl]uracil (1b). 5-Ethyl-6-(2-methylbenzyl)ura-
cil [59] (2.44 g, 10 mmol) was suspended in MeCN (100 ml). BSA (4.48 g, 5.50 ml, 22 mmol) was added in
one portion. The clear soln. was cooled to ꢂ 508, and TMS triflate (6 ml, 30 mmol) was added dropwise.
After stirring at r.t. overnight, TLC showed some starting material present. The soln. was recooled to
ꢂ 508, and another portion of TMS triflate was added (6 ml, 30 mmol). The soln. was allowed to reach r.t.,
and stirring was continued until total conversion of the uracil. After the standard workup [10], the
product was purified by CC (SiO2; CHCl3). Yield: 3.785 g (71%). White solid. Rf (3% MeOH/CHCl3)
0.33. M.p. 121 – 1238. 1H-NMR (CDCl3): 1.06 (t, J ¼ 7.4, MeCH2); 2.38 – 2.47 (m, MeCH2, 2-MeC6H4,
CꢀCH); 4.03 (s, ArCH2); 4.28 (d, J ¼ 2.40, CH2Cꢀ); 5.12 (s, CH2N); 6.83 (d, J ¼ 7.2, 1 arom. H); 7.11 –
7.27 (m, 3 arom. H); 9.74 (s, NH).13C-NMR (CDCl3): 13.70 (MeCH2); 19.04 (MeCH2); 19.66 (2-
MeC6H4); 30.86 (ArCH2); 57.26 (CH2Cꢀ); 72.78 (CH2N); 74.65 (CH2CꢀC); 79.01 (CH2CꢀC); 117.28
(C(5)); 125.55; 126.72; 127.28; 130.59; 133.28; 136.04 (arom. C); 149.28 (C(6)); 152.08 (C(2)); 163.27
(C(4)). HR-MALDI-MS: 335.1363 ([M þ Na]þ, C18H20N2NaO3þ ; calc. 335.1366). Anal. calc. for
C18H20N2O3 · 0.25 H2O (316.88): C 68.23, H 6.52, N 8.84; found: C 68.58, H 6.45, N 8.91.
General Procedure for Sonogashira Coupling for Preparation of 2 – 22. A round-bottomed flask was
charged with the alkyne 1a and 1b (1 mmol), resp., ArI (1.2 mmol), CuI (12 mg, 0.063 mmol), and
[Pd(PPh3)2Cl2] (22 mg, 0.031 mmol). The mixture was flushed with Ar. Dry Et3N (30 ml), previously
bubbled with Ar (25 min), was added. The mixture was stirred at r.t. until the alkyne was consumed (for
16a and 16b, and 17a, the mixture was warmed to 50 – 808). The amine was removed on a rotary
evaporator, and CHCl3 (20 ml) was added. The org. phase was washed with H2O (20 ml). The org. layer
was separated and dried (Na2SO4). The solvent was removed under reduced pressure, and the residue
purified by CC (SiO2; CHCl3).
6-(3,5-Dimethylbenzyl)-5-ethyl-1-{[(3-phenylprop-2-ynyl)oxy]methyl}uracil (2a). Yield: 303 mg
1
(75%). White foam. Rf (CHCl3) 0.14. H-NMR (CDCl3): 1.01 (t, J ¼ 7.4, MeCH2); 2.24 (s, 2 Me); 2.45
(q, J ¼ 7.4, MeCH2); 4.10 (s, ArCH2); 4.51 (s, CH2Cꢀ); 5.26 (s, CH2N); 6.70 (s, 2 arom. H); 6.88 (s, 1
arom. H); 7.25 – 7.41 (m, 5 arom. H); 9.44 (s, NH). 13C-NMR (CDCl3): 13.65 (MeCH2); 19.16 (MeCH2);
21.21 (2 Me); 33.28 (ArCH2); 58.01 (CH2Cꢀ); 72.65 (CH2N); 84.29 (CH2Cꢀ); 86.51 (CH2CꢀC); 116.87
(C(5)); 122.14; 124.98; 128.28; 128.65; 128.97; 131.81; 134.80; 138.88 (arom. C); 149.17 (C(2)); 151.97
(C(6)); 163.26 (C(4)). HR-MALDI-MS: 425.1823 ([M þ Na]þ, C25H26N2NaO3þ ; calc. 425.1836). Anal.
calc. for C25H26N2O3 · 0.25 H2O (407); C 73.78, H 6.56, N 6.88; found: C 73.72, H 6.52, N 6.80.
5-Ethyl-6-(2-methylbenzyl)-1-{[(3-phenylprop-2-ynyl)oxy]methyl}uracil (2b). Yield: 306 mg (79%).
White foam. Rf (3% MeOH/CHCl3) 0.39. 1H-NMR (CDCl3): 0.98 (t, J ¼ 7.4, MeCH2); 2.35 (s, 2-
MeC6H4); 2.39 (q, J ¼ 7.4, MeCH2); 4.05 (s, ArCH2); 4.51 (s, CH2Cꢀ); 5.17 (s, CH2N); 6.82 (d, J ¼ 6.8, 1
arom. H); 7.01 – 7.37 (m, 8 arom. H); 9.56 (s, NH). 13C-NMR (CDCl3): 13.56 (MeCH2); 19.06 (2-
MeC6H4); 19.67 (MeCH2); 30.93 (ArCH2); 58.11 (CH2Cꢀ); 72.93 (CH2N); 84.24 (CH2Cꢀ); 86.52
(CH2CꢀC); 117.16 (C(5)); 122.09; 125.53; 126.70; 127.24; 128.22; 128.60; 130.62; 131.77; 133.36; 136.07
(arom. C); 149.34 (C(2)); 151.98 (C(6)); 163.16 (C(4)). HR-MALDI-MS: 411.1665 ([M þ Na]þ,
C24H24N2NaOþ3 ; calc. 411.1679).
6-(3,5-Dimethylbenzyl)-5-ethyl-1-{[(3-phenanthren-9-ylprop-2-ynyl)oxy]methyl}uracil (3a). Yield:
316 mg (69%). White solid. Rf (3% MeOH/CHCl3) 0.43. M.p. 181 – 1838. 1H-NMR (CDCl3): 0.96 (t, J ¼
7.4, MeCH2); 2.17 (s, 2 Me); 2.42 (q, J ¼ 7.4, MeCH2); 4.13 (s, ArCH2); 4.69 (s, CH2Cꢀ); 5.35 (s, CH2N);
6.68 (s, 2 arom. H); 6.83 (s, 1 arom. H); 7.58 – 7.68 (m, 4 H of phenanthryl); 7.82 – 7.84 (m, 1 H of
phenanthryl); 7.95 (s, 1 H of phenanthryl); 8.36 – 8.39 (m, 1 H of phenanthryl); 8.61 – 8.67 (m, 2 H of
phenanthryl); 9.38 (s, NH). 13C-NMR (CDCl3): 13.64 (MeCH2); 19.17 (MeCH2); 21.16 (2 Me); 33.35
(ArCH2); 58.17 (CH2Cꢀ); 72.68 (CH2N); 84.80 (CH2CꢀC); 88.74 (CH2CꢀC); 116.94 (C(5)); 118.53;

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Helvetica Chimica Acta – Vol. 92 (2009)
122.54; 122.72; 124.96; 126.69; 126.94; 127.07; 127.63; 128.59; 128.96; 130.00; 130.37; 130.97; 132.64;
134.78; 138.87 (arom. C); 149.19 (C(6)); 152.02 (C(2)); 163.21 (C(4)). HR-MALDI-MS: 525.2128 ([M þ
Na]þ, C33H30N2NaO3þ ; calc. 525.2148). Anal. calc. for C33H30N2O3 · 2 H2O (538.65): C 73.59, H 5.61, N
5.20; found: C 73.71, H 5.69, N 5.18.
5-Ethyl-6-(2-methylbenzyl)-1-{[(3-naphthalen-1-ylprop-2-ynyl)oxy]methyl}uracil (4b). Yield:
1
398 mg (83%). White foam. Rf (3% MeOH/CHCl3) 0.36. H-NMR (CDCl3): 0.92 (t, J ¼ 7.4, MeCH2);
2.32 – 2.36 (m, MeCH2, 2-MeC6H4); 4.06 (s, ArCH2); 4.66 (CH2Cꢀ); 5.24 (s, CH2N); 6.81 – 6.84 (m, 1
arom. H); 7.08 – 7.16 (m, 3 arom. H); 7.36 – 7.60 (m, 4 H of naphthyl); 7.77 – 7.87 (m, 2 H of naphthyl);
8.23 – 8.27 (m, 1 H of naphthyl). 13C-NMR (CDCl3): 13.51 (MeCH2); 19.05 (MeCH2); 19.67 (2-MeC6H4);
30.96 (ArCH2); 58.26 (CH2Cꢀ); 72.93 (CH2N); 84.68 (CH2CꢀC); 89.07 (CH2CꢀC); 117.81 (C(5));
119.72; 125.08; 125.51; 125.88; 126.38; 126.69; 126.86; 127.24; 128.24; 129.17; 130.60; 130.91; 133.02;
133.26; 133.32; 136.06 (arom. C); 149.33 (C(2)); 151.94 (C(6)); 162.97 (C(4)). HR-MALDI-MS: 461.1815
([M þ Na]þ, C28H26N2NaO3þ ; calc. 461.1835). Anal. calc. for C28H26N2O3 · 0.75 H2O (452.04): C 74.33, H
5.75, N 6.19; found: C 74.33, H 5.80, N 6.13.
5-Ethyl-1-({[3-(9H-fluoren-2-yl)prop-2-ynyl]oxy}methyl)-6-(2-methylbenzyl)uracil (5b). Yield:
1
454 mg (95%). White foam. Rf (3% MeOH/CHCl3) 0.40. H-NMR (CDCl3): 0.98 (t, J ¼ 7.4, MeCH2);
2.32 – 2.39 (m, MeCH2, 2-MeC6H4); 3.86 (s, CH2); 4.06 (s, ArCH2); 4.53 (s, CH2Cꢀ); 5.19 (s, CH2N); 6.83
(d, J ¼ 6.8, 1 arom. H); 7.10 – 7.76 (m, 10 H of phenyl and fluorenyl); 9.33 (s, NH). 13C-NMR (CDCl3):
13.58 (MeCH2); 19.10 (MeCH2); 19.69 (2-MeC6H4); 30.97 (ArCH2); 36.65 (CH2); 58.21 (CH2Cꢀ); 72.94
(CH2N); 84.10 (CH2CꢀC); 87.30 (CH2CꢀC); 117.16 (C(5)); 119.66; 120.06; 120.21; 125.05; 125.56;
126.72; 126.86; 127.20; 127.24; 128.42; 130.64; 133.40; 136.10; 140.90; 142.20; 143.06; 143.55 (17 arom. C);
149.41 (C(2)); 151.93 (C(6)); 163.08 (C(4)). HR-MALDI-MS: 499.1975 ([M þ Na]þ, C31H28N2NaO3þ ;
calc. 499.1992). Anal. calc. C31H28N2O3 · 1.75 H2O (508.11): C 73.21, H 5.51, N 5.51; found: C 73.29, H
5.58, N 5.47.
1-({[3-(4-Aminophenyl)prop-2-ynyl]oxy}methyl)-6-(3,5-dimethylbenzyl)-5-ethyluracil (6a). Yield:
56 mg (13%). Yellow foam. Rf (3% MeOH/CHCl3) 0.25. 1H-NMR (CDCl3): 1.01 (t, J ¼ 7.4, MeCH2); 2.24
(s, Me); 2.43 (q, J ¼ 7.4, MeCH2); 3.85 (s, NH2); 4.10 (s, ArCH2); 4.48 (s, CH2Cꢀ); 5.23 (s, NCH2O); 6.56
(d, J ¼ 8.6, 2 arom. H); 6.69 (s, 2 arom. H); 6.87 (s, 1 arom. H); 7.19 (d, J ¼ 8.6, 2 arom. H); 9.23 (s, NH).
13C-NMR (CDCl3): 13.66 (MeCH2); 19.16 (MeCH2); 21.21 (2 Me); 32.28 (ArCH2); 58.17 (CH2C ꢀ );
72.56 (CH2N); 81.04 (CH2C ꢀ ); 87.20 (CH2C ꢀ C); 111.37, 114.59 (arom. C); 116.75 (C(5)); 125.01;
128.94; 133.24; 134.83; 138.85; 146.98 (arom. C); 149.32 (C(2)); 151.85 (C(6)); 163.24 (C(4)). HR-
MALDI-MS: 440.1950 ([M þ Na]þ, C25H27N3NaO3þ ; calc. 440.1945).
1-({[3-(4-Aminophenyl)prop-2-ynyl]oxy}methyl)-5-ethyl-6-(2-methylbenzyl)uracil (6b). Yield:
1
80 mg (20%). Yellow foam. Rf (3% MeOH/CHCl3) 0.25. H-NMR (CDCl3): 0.98 (t, J ¼ 7.4, MeCH2);
2.34 (s, Me); 2.34 (q, J ¼ 7.4, MeCH2); 3.83 (s, NH2); 4.05 (s, ArCH2); 4.47 (s, CH2C ꢀ ); 5.15 (s, CH2N);
6.55 (d, J ¼ 8.6, 2 arom. H); 6.82 (d, J ¼ 7.0, 1 arom. H); 7.11 – 7.18 (m, 5 arom. H); 9.25 (s, NH). 13C-NMR
(CDCl3): 13.57 (MeCH2); 19.07 (MeCH2); 19.68 (2-MeC6H4); 30.95 (ArCH2); 58.29 (CH2C ꢀ ); 72.87
(CH2N); 82.00 (CH2C ꢀ ); 87.28 (CH2C ꢀ C); 111.36, 114.56 (arom. C); 117.05 (C(5)); 125.54; 126.68;
127.21; 130.61; 133.21; 133.43; 136.12; 146.95 (arom. C); 149.47 (C(2)); 151.85 (C(6)); 163.07 (C(4)). HR-
MALDI-MS: 426.1773 ([M þ Na]þ, C24H25N3NaOþ3 ; calc. 426.1788). Anal. calc. for C24H25N3O3 · 1.5 H2O
(430.51): C 66.96, H 5.85, N 9.76; found: C 66.89, H 5.92, N 9.55.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-[({3-[(2-methylsulfanyl)phenyl]prop-2-ynyl}oxy)methyl]uracil
(7a). Yield: 372 mg (83%). White solid. Rf (3% MeOH/CHCl3) 0.41. M.p. 166 – 1688. 1H-NMR (CDCl3):
0.99 (t, J ¼ 7.4, MeCH2); 2.22 (s, 2 Me); 2.39 – 2.44 (m, MeS, MeCH2); 4.10 (s, ArCH2); 4.58 (s, CH2C ꢀ );
5.27 (s, CH2N); 6.69 (s, 2 arom. H); 6.86 (s, 1 arom. H); 7.06 – 7.13 (m, 2 arom. H); 7.26 – 7.36 (m, 1 arom.
H); 9.19 (s, NH). 13C-NMR (CDCl3): 13.58 (MeCH2); 14.96 (MeS); 19.17 (MeCH2); 21.19 (2 Me); 33.32
(ArCH2); 57.96 (CH2C ꢀ ); 72.67 (CH2N); 83.93 (CH2C ꢀ C); 90.76 (CH2C ꢀ ); 116.78 (C(5)); 120.13;
123.90; 124.15; 125.01; 128.91; 129.17; 132.73; 134.86; 138.83; 141.88 (arom. C); 149.23 (C(2)); 151.86
(C(6)); 163.16 (C(4)). HR-MALDI-MS: 471.1690 ([M þ Na]þ, C26H28N2NaO3Sþ; calc. 471.1713). Anal.
calc. for C26H25N3O3S · 0.5 H2O (457.60): C 68.25, H 6.17, N 6.12; found: C 68.07, H 6.14, N 5.98.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-({[3-(2-nitrophenyl)prop-2-ynyl]oxy}methyl)uracil (8a). Yield:
1
288 mg (64%). Brown foam. Rf (3% MeOH/CHCl3) 0.40. H-NMR (CDCl3): 1.00 (t, J ¼ 7.4, MeCH2);
2.25 (s, 2 Me); 2.43 (q, J ¼ 7.4, MeCH2); 4.11 (s, ArCH2); 4.57 (s, CH2C ꢀ ); 5.30 (s, CH2N); 6.71 (s, 2

Helvetica Chimica Acta – Vol. 92 (2009)
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arom. H); 6.88 (s, 1 arom. H); 7.43 – 7.59 (m, 3 arom. H); 8.02 (d, J ¼ 8.0, 1 arom. H); 9.61 (s, NH).
13C-NMR (CDCl3): 13.57 (MeCH2); 18.96 (MeCH2); 21.04 (2 Me); 33.14 (ArCH2); 57.76 (CH2C ꢀ );
72.52 (CH2N); 81.46 (CH2C ꢀ C); 92.28 (CH2C ꢀ ); 116.90 (C(5)); 113.50; 124.55; 124.97; 128.95; 129.04;
132.86; 134.77; 134.93; 138.85 (arom. C); 149.22 (C(2)); 149.60 (arom. C); 152.03 (C(6)); 163.31 (C(4)).
HR-MALDI-MS: 470.1709 ([M þ Na]þ, C25H25N3NaOþ5 ; calc. 470.1686).
6-(3,5-Dimethylbenzyl)-5-ethyl-1-({[3-(4-nitrophenyl)prop-2-ynyl]oxy}methyl)uracil (9a). Yield:
1
358 mg (80%). Yellowish solid. Rf (3% MeOH/CHCl3) 0.41. M.p. 144 – 1468. H-NMR (CDCl3): 1.03
(t, J ¼ 7.4, MeCH2); 2.27 (s, 2 Me); 2.46 (q, J ¼ 7.4, MeCH2); 4.10 (s, ArCH2); 4.55 (s, CH2C ꢀ ); 5.28 (s,
CH2N); 6.71 (s, 2 arom. H); 6.90 (s, 1 arom. H); 7.53 (d, J ¼ 8.9, 2 arom. H); 8.15 (d, J ¼ 8.9, 2 arom. H);
9.53 (s, NH). 13C-NMR (CDCl3): 13.72 (MeCH2); 19.16 (MeCH2); 21.25 (2 Me); 33.33 (ArCH2); 57.85
(CH2C ꢀ ); 72.69 (CH2N); 84.52 (CH2C ꢀ ); 89.69 (CH2C ꢀ C); 117.06 (C(5)); 123.54; 124.95; 128.96;
129.08; 132.54; 134.66; 138.98; 147.31 (arom. C); 149.12 (C(2)); 152.06 (C(6)); 163.23 (C(4)). HR-
MALDI-MS: 470.1673 ([M þ Na]þ, C25H25N3NaO5þ ; calc. 470.1686). Anal. calc. for C25H25N3O5 (447.50):
C 67.10, H 5.63, N 9.39; found: C 66.86, H 5.52, N 9.15.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-[({3-[2-(trifluoromethyl)phenyl]prop-2-ynyl}oxy)methyl]uracil
(10a). Yield: 153 mg (65%). White solid. Rf (3% MeOH/CHCl3) 0.38. M.p. 129 – 1318. 1H-NMR
(CDCl3): 0.99 (t, J ¼ 7.4, MeCH2); 2.24 (s, 2 Me); 2.42 (q, J ¼ 7.4, MeCH2); 4.09 (s, ArCH2); 4.55 (s,
CH2C ꢀ ); 5.26 (s, CH2N); 6.69 (s, 2 arom. H); 6.88 (s, 1 arom. H); 7.41 – 7.65 (m, 4 arom. H); 9.15 (s, NH).
13C-NMR (CDCl3): 13.53 (MeCH2); 19.17 (MeCH2); 21.21 (2 Me); 33.25 (ArCH2); 57.91 (CH2C ꢀ );
72.72 (CH2N); 82.33 (CH2C ꢀ C); 90.07 (CH2C ꢀ ); 116.85 (C(5)); 121.55; 124.98; 125.79 (q, J ¼ 5.0,
CF3); 128.43; 128.98; 131.45; 134.22; 134.78; 138.89 (arom. C); 149.23 (C(2)); 151.88 (C(6)); 163.13
(C(4)). HR-MALDI-MS: 493.1690 ([M þ Na]þ, C26H25F3N2NaOþ3 ; calc. 493.1710). Anal. calc. for
C26H25F3N2O3 (470.50): C 66.37, H 5.36, N5.95; found: C 66.22, H 5.35, N 5.86.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-({[3-(3-cyanophenyl)prop-2-ynyl]oxy}methyl)uracil (11a). Yield:
1
272 mg (64%). White foam. Rf (3% MeOH/CHCl3) 0.33. H-NMR (CDCl3): 1.02 (t, J ¼ 7.4, MeCH2);
2.26 (s, 2 Me); 2.45 (q, J ¼ 7.4, MeCH2); 4.09 (s, ArCH2); 4.52 (s, CH2C ꢀ ); 5.27 (s, CH2N); 6.70 (s, 2
arom. H); 6.90 (s, 1 arom. H); 7.39 – 7.44 (m, 1 arom. H); 7.57 – 7.58 (m, 3 arom. H); 9.55 (s, NH).
13C-NMR (CDCl3): 13.68 (MeCH2); 19.15 (MeCH2); 21.24 (2 Me); 33.32 (ArCH2); 57.78 (CH2C ꢀ );
72.64 (CH2N); 84.04 (CH2C ꢀ ); 86.97 (CH2C ꢀ C); 112.88 (arom. C); 117.02 (C(5)); 117.82 (CN); 123.78;
124.95; 129.06; 129.26; 131.86; 134.69; 135.11; 135.82; 138.94 (arom. C); 149.12 (C(2)); 152.06 (C(6));
163.26 (C(4)). HR-MALDI-MS: 450.1773 ([M þ Na]þ, C26H25N3NaO3þ ; calc. 450.1788). Anal. calc. for
C26H25N3O3 · 1.0 H2O (445.52): C 70.10, H 5.66, N 9.43; found: C 70.40, H 5.61, N 9.29.
1-[({3-[3,5-Bis(trifluoromethyl)phenyl]prop-2-ynyl}oxy)methyl]-6-(3,5-dimethylbenzyl)-5-ethylura-
cil (12a). Yield: 460 mg (86%). White solid. Rf (3% MeOH/CHCl3) 0.38. M.p. 153 – 1558. 1H-NMR
(CDCl3): 1.02 (t, J ¼ 7.4, MeCH2); 2.26 (s, 2 Me); 2.46 (q, J ¼ 7.4, MeCH2); 4.09 (s, ArCH2); 4.53 (s,
CH2C ꢀ ); 5.27 (s, CH2N); 6.70 (s, 2 arom. H); 6.90 (s, 1 arom. H); 7.81 (s, 1 arom. H); 7.85 (s, 2 arom. H);
9.59 (s, NH). 13C-NMR (CDCl3): 13.68 (MeCH2); 19.14 (MeCH2); 21.22 (2 Me); 33.35 (ArCH2);
57.71(CH2C ꢀ ); 72.60 (CH2N); 83.37 (CH2C ꢀ C); 88.05 (OCH2C ꢀ ); 117.13 (C(5)); 122.10; 124.52;
124.60; 124.94; 129.09; 131.72; 132.16; 132.21; 134.65; 138.99 (arom. C); 149.02 (C(6)); 152.12 (C(2));
163.23 (C(4)). HR-MALDI-MS: 561.1582 ([M þ Na]þ, C27H24F6N2NaOþ3 ; calc. 561.1583). Anal. calc. for
C27H24F6N2O3 (538.49): C 60.22, H 4.49, N 5.20; found: C 60.12, H 4.35, N 5.14.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-{[(3-thien-2-ylprop-2-ynyl)oxy]methyl}uracil (13a). Yield: 204 mg
1
(51%). White foam. Rf (3% MeOH/CHCl3) 0.34. H-NMR (CDCl3): 1.00 (t, J ¼ 7.4, MeCH2); 2.24 (s,
2 Me); 2.45 (q, J ¼ 7.4, MeCH2); 4.09 (s, ArCH2); 4.53 (s, CH2C ꢀ ); 5.25 (s, CH2N); 6.70 (s, 2 arom. H);
6.88 (s, 1 arom. H); 6.94 (dd, J ¼ 5.2, 3.6, 1 H of thienyl); 7.18 (dd, J ¼ 3.6, 1.1, 1 H of thienyl); 7.24 (dd, J ¼
5.1, 1.1, 1 H of thienyl); 9.68 (s, NH). 13C-NMR (CDCl3): 13.60 (MeCH2); 19.13 (MeCH2); 21.20 (2 Me);
33.26 (ArCH2); 58.14 (CH2C ꢀ ); 72.74 (CH2N); 79.70 (CH2C ꢀ C); 88.44 (CH2C ꢀ ); 116.96 (C(5));
121.98; 124.98; 126.94; 127.66; 128.96; 132.76; 134.77; 138.87 (arom. C); 149.12 (C(2)); 152.08 (C(6));
163.38 (C(4)). HR-MALDI-MS: 431.1396 ([M þ Na]þ, C23H24N2NaO3Sþ; calc. 431.1399). Anal. calc. for
C23H24N2O3S · 0.5 H2O (417.53): C 66.16, H 5.79, N 6.71; found: C 66.31, H 5.90, N 6.65.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-{[(3-pyridin-3-ylprop-2-ynyl)oxy]methyl}uracil (14a). Yield:
304 mg (75%). White solid. Rf (3% MeOH/CHCl3) 0.28. M.p. 146 – 1488. 1H-NMR (CDCl3): 1.02 (t,
J ¼ 7.4, MeCH2); 2.25 (s, 2 Me); 2.45 (q, J ¼ 7.4, MeCH2); 4.09 (s, ArCH2); 4.53 (CH2C ꢀ ); 5.27 (s,

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CH2N); 6.70 (s, 2 arom. H); 6.89 (s, 1 arom. H); 7.20 – 7.30 (m, 1 H of pyridinyl); 7.68 (d, J ¼ 8.0, 1 H of
pyridinyl); 8.48 – 8.80 (m, 2 H of pyridinyl); 9.88 (s, NH). 13C-NMR (CDCl3): 13.68 (MeCH2); 19.15
(MeCH2); 21.22 (2 Me); 33.30 (ArCH2); 57.87 (CH2C ꢀ ); 72.65 (CH2N); 83.12 (CH2C ꢀ C); 87.88
(CH2C ꢀ ); 116.99 (C(5)); 124.95; 129.01; 134.74; 138.75; 138.91; 149.03; 152.10 (arom. C); 148.85 (C(2));
152.32 (C(6)); 163.34 (C(4)). HR-MALDI-MS: 426.1770 ([M þ Na]þ, C24H25N3NaO3þ ; calc. 426.1788).
Anal. calc. for C24H25N3O3 · 0.25 H2O (407.99): C 70.66, H 6.18, N 10.30; found: C 70.48, H 6.18, N 10.13.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-{[(3-pyrazin-2-ylprop-2-ynyl)oxy]methyl}uracil (15a). Yield:
1
230 mg (56%). White foam. Rf (3% MeOH/CHCl3) 0.24. H-NMR (CDCl3): 1.02 (t, J ¼ 7.4, MeCH2);
2.26 (s, 2 Me); 2.45 (q, J ¼ 7.4, MeCH2); 4.09 (s, ArCH2); 4.57 (CH2C ꢀ ); 5.27 (s, CH2N); 6.71 (s, 2 arom.
H); 6.89 (s, 1 arom. H); 8.50 – 8.64 (m, 3 H of pyrazinyl); 9.46 (s, NH). 13C-NMR (CDCl3): 13.68
(MeCH2); 19.17 (MeCH2); 21.23 (2 Me); 33.31 (ArCH2); 57.74 (CH2C ꢀ ); 72.81 (CH2N); 82.89
(CH2C ꢀ C); 88.63 (CH2C ꢀ C); 117.01 (C(5)); 124.96; 129.04; 134.70; 138.93; 143.33; 144.45; 147.63
(arom. C); 148.93 (C(2)); 152.02 (C(6)); 163.19 (C(4)). HR-MALDI-MS: 427.1738 ([M þ Na]þ,
C23H24N4NaOþ3 ; calc. 427.1741). Anal. calc. for C23H24N4O3 · 0.5 H2O (413.48): C 66.81, H 5.85, N 13.55;
found: C 66.59, H 5.84, N 13.37.
N-[4-(3-{[6-(3,5-Dimethylbenzyl)-5-ethyl-3,4-dihydro-2,4-dioxo-2H-pyrimidin-1-yl]methoxy}prop-
1-ynyl)phenyl]-2,2,2-trifluoroacetamide (16a). Yield: 830 mg (81%). Yellow foam. Rf (3% MeOH/
CHCl3) 0.25. 1H-NMR (CDCl3): 0.98 (t, J ¼ 7.4, MeCH2); 2.25 (s, 2 Me); 2.41 (q, J ¼ 7.4, MeCH2); 4.10 (s,
ArCH2); 4.49 (s, CH2Cꢀ); 5.26 (s, CH2N); 6.69 (s, 2 arom. H); 6.89 (s, 1 arom. H); 7.36 (d, J ¼ 8.7, 2 arom.
H); 7.55 (d, J ¼ 8.7, 2 arom. H); 8.53 (s, NH); 9.60 (s, NH). 13C-NMR (CDCl3): 13.65 (MeCH2); 19.12
(MeCH2); 21.23 (2 Me); 33.32 (ArCH2); 58.05 (CH2C ꢀ ); 72.79 (CH2N); 85.00 (CH2C ꢀ );
85.61(CH2C ꢀ C); 116.96 (C(5)); 117.52; 119.87; 120.31; 124.97; 129.05; 132.75; 134.71; 135.62; 138.95
(arom. C); 149.49 (C(2)); 152.09 (C(6)); 163.46 (C(4)). HR-MALDI-MS: 536.1749 ([M þ Na]þ,
C27H26F3N3NaOþ4 ; calc. 536.1768).
N-[4-(3-{[5-Ethyl-6-(2-methylbenzyl)-3,4-dihydro-2,4-dioxo-2H-pyrimidin-1-yl]methoxy}prop-1-
ynyl)phenyl]-2,2,2-trifluoroacetamide (16b). Yield: 407 mg (81%). Yellow foam. Rf (3% MeOH/CHCl3)
1
0.25. H-NMR (CDCl3): 0.95 (t, J ¼ 7.4, MeCH2); 2.32 – 2.39 (m, MeCH2, 2-MeC6H4); 4.05 (s, ArCH2);
4.48 (s, CH2Cꢀ); 5.17 (s, CH2N); 6.82 (d, J ¼ 6.9, 2 arom. H); 7.10 – 7.20 (m, 3 arom. H); 7.33 (d, J ¼ 8.7, 2
arom. H); 7.54 (d, J ¼ 8.7, 2 arom. H); 8.57 (s, NH); 9.68 (s, NH). 13C-NMR (CDCl3): 13.58 (MeCH2);
19.03 (MeCH2); 19.68 (2-MeC6H4); 30.92 (ArCH2); 58.11 (CH2C ꢀ ); 73.04 (CH2N); 84.96 (CH2C ꢀ );
85.65 (CH2C ꢀ C); 117.24 (C(5)); 119.82; 120.30; 125.53; 126.77; 127.33; 132.73; 135.63; 136.06 (arom. C);
149.67 (C(2)); 152.09 (C(6)); 163.35 (C(4)). HR-MALDI-MS: 522.1607 ([M þ Na]þ, C26H24F3N3NaO4þ ;
calc. 522.1611).
2-Bromo-N-[4-(3-{[6-(3,5-dimethylbenzyl)-5-ethyl-3,4-dihydro-2,4-dioxo-2H-pyrimidin-1-yl]me-
thoxy}prop-1-ynyl)phenyl]acetamide (17a). Compound 1a (163 mg, 0.5 mmol), 2-bromo-N-(4-iodophe-
nyl)acetamide [58] (204 mg, 0.6 mmol), CuI (12 mg, 0.062 mmol), and [PdCl2(PPh3)2] (22 mg,
0.031 mmol) were dissolved in dry THF/Et3N 3 :1 (20 ml). The resulting soln. was stirred at 508 under
Ar until complete consumption of 1a. Solvents were removed, and the residue was submitted to CC
(SiO2; 5% MeOH/CHCl3). Yield: 4 mg (1.5%).
1-[({6-[(tert-Butyl)diphenylsilyloxy]hexa-2,4-diynyl}oxy)methyl]-6-(3,5-dimethylbenzyl)-5-ethyl-
1H-pyrimidine-2,4-dione (18a). Yield: 293 mg (47%). Brown foam. Rf (3% MeOH/CHCl3) 0.51; also
193 mg of homocoupled product [21] was isolated. Rf (3% MeOH/CHCl3) 0.31. 1H-NMR (CDCl3): 1.05
t
(s, Bu); 1.08 (t, J ¼ 7.4, MeCH2); 2.28 (s, 2 Me); 2.47 (q, J ¼ 7.4, MeCH2); 4.05 (s, ArCH2); 4.34 (s,
CH2Cꢀ); 4.35 (s, CH2OSi); 5.16 (s, CH2N); 6.71 (s, 2 arom. H); 6.90 (s, 1 arom. H); 7.36 – 7.46 (m, 6 arom.
H); 7.67 (d, J ¼ 7.6, 4 arom. H); 9.05 (s, NH). 13C-NMR (CDCl3): 13.77 (MeCH2); 19.15 (Me3C); 19.22
(MeCH2); 21.29 (2 Me); 26.61 (Me3C); 33.30 (ArCH2); 52.93 (CH2OSi); 57.74 (CH2C ꢀ ); 68.72
(CH2C ꢀ CꢂC ꢀ C); 70.87 (CH2C ꢀ CꢂC ꢀ C); 72.64 (CH2N); 74.17 (CH2C ꢀ CꢂC ꢀ C); 78.26
(CH2C ꢀ CꢂC ꢀ C); 117.05 (C(5)); 125.02; 127.78; 129.06; 129.89; 132.60; 134.69; 135.53; 138.96 (arom.
C); 148.94 (C(6)); 151.84 (C(2)); 163.01 (C(4)). HR-MALDI-MS: 641.2801 ([M þ Na]þ,
C38H42N2NaO4Siþ; calc. 641.2806). Anal. calc. for C38H42N2O4Si · 0.5 H2O (627.86): C 72.69, H 6.90, N
4.46; found: C 72.86, H 6.82, N 4.26.
1-{[(3-{4-[(tert-Butyl)diphenylsilyloxy]phenyl}prop-2-ynyl)oxy]methyl}-6-(3,5-dimethylbenzyl)-5-
1
ethyluracil (19a). Yield: 625 mg (69%). White foam. Rf (3% MeOH/CHCl3) 0.38. H-NMR (CDCl3):

Helvetica Chimica Acta – Vol. 92 (2009)
1397
t
0.99 (t, J ¼ 7.4, MeCH2); 1.09 (s, Bu); 2.21 (s, 2 Me); 2.42 (q, J ¼ 7.4, MeCH2); 4.06 (s, ArCH2); 4.44 (s,
OCH2Cꢀ); 5.20 (s, CH2N); 6.66 (s, 2 arom. H); 6.68 (d, J ¼ 8.7, 2 arom. H); 6.85 (s, 1 arom. H); 7.13 (d,
J ¼ 8.7, 2 arom. H); 7.32 – 7.42 (m, 6 arom. H); 7.68 (d, J ¼ 7.8, 4 arom. H); 8.94 (s, NH). 13C-NMR
(CDCl3): 13.66 (MeCH2); 19.18 (MeCH2); 19.40 (Me3C); 21.22 (2 Me); 26.40 (Me3C); 33.27 (ArCH2);
57.94 (CH2C ꢀ ); 72.48 (CH2N); 82.90 (CH2C ꢀ ); 86.62 (CH2C ꢀ C); 114.60 (arom. C); 116.77 (C(5));
119.82; 124.98; 127.81; 128.96; 130.00; 132.39; 133.13; 134.79; 135.40; 138.87 (arom. C); 149.20 (C(2));
151.74 (C(6)); 156.11 (arom. C); 163.02 (C(4)). HR-MALDI-MS: 679.2952 ([Mþ Na]þ, C41H44N2NaO4Siþ;
calc. 679.2963). Anal. calc. for C41H44N2O4Si · 0.25 H2O (661.41): C 74.46, H 6.79, N 4.24; found: C 74.35,
H 6.69, N 4.27.
1-{[(3-{3-[(tert-Butyl)diphenylsilyloxy]phenyl}prop-2-ynyl)oxy]methyl}-6-(3,5-dimethylbenzyl)-5-
1
ethyluracil (20a). Yield: 426 mg (65%). White foam. Rf (3% MeOH/CHCl3) 0.46. H-NMR (CDCl3):
t
1.02 (t, J ¼ 7.4, MeCH2); 1.08 (s, Bu); 2.23 (s, 2 Me); 2.44 (q, J ¼ 7.4, MeCH2); 4.06 (s, ArCH2); 4.44 (s,
OCH2Cꢀ); 5.19 (s, CH2N); 6.63 (ddd, J ¼ 8.0, 2.4, 1.5, 1 arom. H); 6.67 (s, 2 arom. H); 6.87 (s, 1 arom.
H); 6.91 – 6.93 (m, 2 arom. H); 6.96 (t, J ¼ 7.9, 1 arom. H); 7.32 – 7.41 (m, 6 arom. H); 7.68 (d, J ¼ 7.8, 4
arom. H); 8.89 (s, NH). 13C-NMR (CDCl3): 13.69 (MeCH2); 19.21 (MeCH2); 19.41 (Me3C); 21.22
(2 Me); 26.42 (Me3C); 33.29 (ArCH2); 57.88 (CH2C ꢀ ); 72.53 (CH2N); 83.94 (CH2C ꢀ ); 86.35 (CH2C ꢀ
C); 116.81 (C(5)); 120.49; 122.99; 124.84; 125.01; 127.80; 128.99; 129.12; 129.95; 132.51; 134.76; 135.44;
138.89 (arom. C); 149.16 (C(2)); 151.74 (C(6)); 155.31 (arom. C); 162.99 (C(4)). HR-MALDI-MS:
679.2957 ([M þ Na]þ, C41H44N2NaO4Siþ; calc. 679.2963). Anal. calc. for C41H44N2O4Si · 0.25 H2O
(661.41): C 74.46, H 6.79, N 4.24; found: C 74.40, H 6.68, N 4.27.
1-({[3-(4-{[(tert-Butyl)diphenylsilyloxy]methyl}phenyl)prop-2-ynyl]oxy}methyl)-6-(3,5-dimethyl-
benzyl)-5-ethyluracil (21a). Yield: 277 mg (41%). White foam. Rf (CHCl3) 0.47. 1H-NMR (CDCl3): 1.02
t
(t, J ¼ 7.4, MeCH2); 1.09 (s, Bu); 2.23 (s, 2 Me); 2.44 (q, J ¼ 7.4, MeCH2); 4.10 (s, ArCH2); 4.51 (s,
CH2OSi); 4.74 (s, CH2C ꢀ ); 5.25 (s, CH2N); 6.70 (s, 2 arom. H); 6.88 (s, 1 arom. H); 7.27 (d, J ¼ 8.1, 2
arom. H); 7.34 – 7.42 (m, 8 arom. H); 7.67 (d, J ¼ 7.9, 4 arom. H); 9.17 (s, NH). 13C-NMR (CDCl3): 13.68
(MeCH2); 19.20 (MeCH2); 19.28 (Me3C); 21.23 (2 Me); 26.79 (Me3C); 33.31 (ArCH2); 57.98 (CH2C ꢀ );
65.14 (CH2OSi); 72.58 (CH2N); 83.85 (CH2C ꢀ ); 86.68 (CH2C ꢀ C); 116.85 (C(5)); 120.55; 125.00;
125.82; 127.72; 128.99; 129.73; 131.74; 133.27; 134.80; 135.49; 138.89; 141.82 (arom. C); 149.19 (C(2));
151.86 (C(6)); 163.13 (C(4)). HR-MALDI-MS: 693.3113 ([M þ Na]þ, C42H46N2NaO4Siþ; calc. 693.3119).
Anal. calc. for C42H46N2O4Si · 0.25 H2O (675.44): C 74.69, H 6.93, N 4.15; found: C 74.60, H 6.94, N 4.11.
1-({[3-(3-{[(tert-Butyl)diphenylsilyloxy]methyl}phenyl)prop-2-ynyl]oxy}methyl)-6-(3,5-dimethyl-
benzyl)uracil (22a). Yield: 315 mg (47%). White foam. Rf (CHCl3) 0.51. 1H-NMR (CDCl3): 1.01 (t, J ¼
7.4, MeCH2); 1.09 (s, tBu); 2.22 (s, 2 Me); 2.43 (q, J ¼ 7.4, MeCH2); 4.09 (s, ArCH2); 4.50 (s, CH2OSi); 4.72
(s, CH2C ꢀ ); 5.24 (s, CH2N); 6.69 (s, 2 arom. H); 6.86 (s, 1 arom. H); 7.28 – 7.42 (m, 10 arom. H); 7.66 –
7.67 (d, J ¼ 7.6, 4 arom. H); 8.98 (s, NH). 13C-NMR (CDCl3): 13.67 (MeCH2); 19.20 (MeCH2); 19.28
(Me3C); 21.21 (2 Me); 26.82 (Me3C); 33.31 (ArCH2); 57.97 (CH2C ꢀ ); 65.00 (CH2OSi); 72.57 (CH2N);
84.01 (CH2C ꢀ ); 86.74 (CH2C ꢀ C); 116.83 (C(5)); 121.95; 125.00; 126.43; 127.73; 128.26; 128.99; 129.29;
129.73; 130.42; 133.27; 134.78; 135.52; 141.26 (arom. C); 149.19 (C(2)); 151.78 (C(6)); 163.04 (C(4)). HR-
MALDI-MS: 693.3095 ([M þ Na]þ, C42H46N2NaO4Siþ; calc. 693.3119). Anal. calc. for C42H46N2O4Si ·
0.25 H2O (675.44): C 74.69, H 6.93, N 4.15; found: C 74.53, H 6.94, N 4.19.
General Procedure for the Synthesis of 17a and 17b from 6a and 6b, Resp. Deprotection of Amides 16a
and 16b. Powdered KOH (170 mg, 3 mmol) was dissolved in iPrOH (20 ml), and 16a or 16b (1 mmol) was
added to the soln. The soln. was heated at 908 (oil bath) for 2 – 3 h. After cooling, the mixture was
evaporated to dryness, and H2O (20 ml) was added. The mixture was extracted with CHCl3 (2 ꢃ 20 ml).
After drying (Na2SO4), the solvent was removed under reduced pressure, and the residue was purified by
CC (SiO2; CHCl3) to deliver amines 6a or 6b, resp. Yields 87 and 82%, resp. The amines were used for the
next step.
Conversion of 6a and 6b to 17a and 17b, Resp. Compounds 6a and 6b (1 mmol) were dissolved in dry
THF (10 ml), and (BrCH2CO)2O (520 mg, 2 mmol) dissolved in dry THF (5 ml) was added dropwise at
08. The mixture was further stirred for 2 h at r.t. After evaporation of THF, the residue was dissolved in
AcOEt (20 ml) and washed with sat. aq. soln. of NaHCO3 and sat. aq. soln. of NaCl, dried (Na2SO4), and
concentrated.

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Helvetica Chimica Acta – Vol. 92 (2009)
2-Bromo-N-[4-(3-{[6-(3,5-dimethylbenzyl)-5-ethyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl]meth-
oxy}prop-1-ynyl)phenyl]acetamide (17a). Yield: 484 mg (90%). Light brown foam. Rf (5% MeOH/
CHCl3) 0.36. 1H-NMR (CDCl3): 0.99 (t, J ¼ 7.5, MeCH2); 2.25 (s, 2 Me); 2.42 (q, J ¼ 7.5, MeCH2); 4.01 (s,
ArCH2); 4.10 (s, NHCOCH2); 4.49 (s, CH2C ꢀ ); 5.26 (s, CH2N); 6.69 (s, 2 arom. H); 6.88 (s, 1 arom. H);
7.36 (d, J ¼ 8.7, 2 arom. H); 7.50 (d, J ¼ 8.7, 2 arom. H); 8.31 (s, NH); 9.65 (s, NH). 13C-NMR (CDCl3):
13.64 (MeCH2); 19.12 (MeCH2); 21.24 (2 Me); 30.29 (CH2Br); 33.32 (ArCH2); 58.13 (CH2C ꢀ ); 72.77
(CH2N); 84.33 (CH2C ꢀ ); 85.98 (CH2C ꢀ C); 116.79 (C(5)); 119.55; 124.96; 129.05; 132.67; 134.64;
137.32; 138.94 (arom. C). 149.80 C(2); 151.94 (C(6)); 163.73 (C(4)); 170.81 (COCH2Br). HR-MALDI-
MS: 560.1137 ([M þ Na]þ, C27H28BrN3NaOþ4 ; calc. 560.1155).
2-Bromo-N-[4-(3-{[5-ethyl-6-(2-methylbenzyl)-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl]methoxy}-
prop-1-ynyl)phenyl]acetamide (17b). Yield: 361 mg (69%). Light brown solid. Rf (5% MeOH/CHCl3)
1
0.32. M.p. 162 – 1638. H-NMR (CDCl3): 0.98 (t, J ¼ 7.5, MeCH2); 2.34 – 2.41 (m, Me, MeCH2); 4.02 (s,
ArCH2); 4.05 (s, NHCOCH2); 4.49 (s, CH2C ꢀ ); 5.15 (s, CH2N); 6.82 (d, J ¼ 7.2, 1 arom. H); 7.10 – 7.19
(m, 3 arom. H); 7.33 (d, J ¼ 8.7, 2 arom. H); 7.48 (d, J ¼ 8.7, 2 arom. H); 8.31 (s, NH); 9.56 (s, NH).
13C-NMR (CDCl3): 13.58 (MeCH2); 19.05 (MeCH2); 19.70 (2-MeC6H4); 29.37 (ArCH2); 30.95 (CH2Br);
58.18 (CH2C ꢀ ); 73.03 (CH2N); 84.29 (CH2C ꢀ ); 86.01 (CH2C ꢀ C); 117.09 (C(5)); 119.53; 125.51;
126.75; 127.33; 130.67; 132.65; 133.23; 136.08; 137.31 (arom. C); 149.88 (C(2)); 151.91 (C(6)); 163.47
(C(4)); 171.37 (COCH2Br). HR-MALDI-MS: 546.0988 ([M þ Na]þ, C26H26BrN3NaO4þ ; calc. 546.0999).
General Procedure for the Deprotection of the Silyl Alcohols. The appropriate alcohol 18a – 22a (0.5 –
1.5 mmol) was treated with TBAF (1m in THF, 0.5 – 1.5 ml) in THF (20 ml) at r.t. until TLC showed no
more starting material. H2O (15 ml) was added, and the mixture was extracted with AcOEt (2 ꢃ 15 ml).
The org. phase was dried (Na2SO4) and evaporated under reduced pressure. The residue was purified by
CC (SiO2; 5% AcOEt/CHCl3).
6-(3,5-Dimethylbenzyl)-5-ethyl-1-{[(6-hydroxyhexa-2,4-diynyl)oxy]methyl}uracil (23a). Yield:
1
101 mg (48%). White foam. Rf (3% MeOH/CHCl3) 0.24. H-NMR (CDCl3): 1.12 (t, J ¼ 7.4, MeCH2);
2.29 (s, 2 Me); 2.51 (q, J ¼ 7.4, MeCH2); 3.39 (br. s, OH); 4.10 (s, ArCH2); 4.26 (d, J ¼ 2.7, ꢀ CH2OH);
4.35 (s, CH2C ꢀ ); 5.29 (s, CH2N); 6.72 (s, 2 arom. H); 6.90 (s, 1 arom. H); 9.76 (s, NH). 13C-NMR
(CDCl3): 13.79 (MeCH2); 19.14 (MeCH2); 21.27 (2 Me); 33.18 (ArCH2); 51.05 (CH2OH); 58.37
(OCH2C ꢀ ); 69.11 (OCH2C ꢀ CꢂC ꢀ C); 70.35 (OCH2C ꢀ CꢂC ꢀ C); 73.48 (OCH2C ꢀ CꢂC ꢀ C);
75.47 (CH2N); 79.64 (OCH2C ꢀ CꢂC ꢀ C); 117.57 (C(5)); 125.08; 129.07; 134.75; 138.96 (arom. C);
149.72 (C(6)); 152.49 (C(2)); 164.14 (C(4)). HR-MALDI-MS: 403.1616 ([M þ Na]þ, C22H24N2NaO4þ ;
calc. 403.1628). Anal. calc. for C22H24N2O4 · 1.0 H2O (398.46): C 66.32, H 6.58, N 7.03; found: C 66.54, H
6.12, N 7.03.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-({[3-(4-hydroxyphenyl)prop-2-ynyl]oxy}methyl)uracil (24a).
Yield: 300 mg (81%). White foam. Rf (3% MeOH/CHCl3) 0.18. 1H-NMR (CDCl3): 0.96 (t, J ¼ 7.4,
MeCH2); 2.24 (s, 2 Me); 2.42 (q, J ¼ 7.4, MeCH2); 4.11 (s, ArCH2); 4.48 (s, CH2C ꢀ ); 5.27 (s, CH2N); 6.69
(s, 2 arom. H); 6.77 (d, J ¼ 8.7, 2 arom. H); 6.88 (s, 1 arom. H); 7.00 (br. s, OH); 7.22 (d, J ¼ 8.7, 2 arom. H);
9.60 (s, NH). 13C-NMR (CDCl3): 13.60 (MeCH2); 19.15 (MeCH2); 21.24 (2 Me); 33.31 (ArCH2); 58.34
(CH2C ꢀ ); 72.91 (CH2N); 82.67 (CH2C ꢀ ); 86.69 (CH2C ꢀ C); 113.66 (arom. C); 115.59 (C(5)); 116.92;
125.02; 129.03; 133.47; 134.73; 138.93 (arom. C); 149.87 (C(2)); 152.14 (C(6)); 156.75 (arom. C); 163.65
(C(4)). HR-MALDI-MS: 441.1772 ([M þ Na]þ, C25H26N2NaO4þ ; calc. 441.1785). Anal. calc. for
C25H26N2O4 · 2 H2O (454.53): C 66.06, H 5.77, N 6.16; found: C 66.16, H 5.77, N 6.06.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-({[3-(3-hydroxyphenyl)-prop-2-ynyl]oxy}methyl)uracil (25a).
Yield: 217 mg (85%). White foam. Rf (3% MeOH/CHCl3) 0.20. 1H-NMR (CDCl3): 1.03 (t, J ¼ 7.4,
MeCH2); 2.27 (s, 2 Me); 2.45 (q, J ¼ 7.4, MeCH2); 4.12 (s, ArCH2); 4.44 (s, CH2C ꢀ ); 5.38 (s, CH2N); 6.69
(s, 2 arom. H); 6.83 – 6.91 (m, 3 arom. H); 7.13 (t, J ¼ 7.9, 1 arom. H); 7.24 (br. s, OH), 7.66 (s, 1 arom. H);
10.88 (s, NH). 13C-NMR (CDCl3): 13.55 (MeCH2); 19.23 (MeCH2); 21.26 (2 Me); 33.33 (ArCH2); 57.04
(CH2C ꢀ ); 71.61 (CH2N); 83.17 (CH2C ꢀ ); 87.64 (CH2C ꢀ C); 116.61 (C(5)); 117.23; 119.43; 122.75;
122.90; 124.96; 129.10; 129.50; 134.66; 139.02 (arom. C); 150.31 (C(6)); 152.88 (C(2)); 156.51 (arom. C);
164.13 (C(4)). HR-MALDI-MS: 441.1774 ([M þ Na]þ, C25H26N2NaO4þ ; 441.1785). Anal. calc. for
C25H26N2O4 · 1.5 H2O (445.52): C 67.40, H 5.88, N 6.29; found: C 6.94, H 5.81, N 6.27.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-[({3-[4-(hydroxymethyl)phenyl]prop-2-ynyl}oxy)methyl]uracil
(26a). Yield: 136 mg (84%). White foam. Rf (3% MeOH/CHCl3) 0.18. 1H-NMR (CDCl3): 0.99 (t, J ¼ 7.4,

Helvetica Chimica Acta – Vol. 92 (2009)
1399
MeCH2); 2.24 (s, 2 Me); 2.43 (q, J ¼ 7.4, MeCH2); 4.10 (s, ArCH2); 4.50 (s, CH2C ꢀ ); 4.66 (s, CH2OH);
5.26 (s, CH2N); 6.70 (s, 2 arom. H); 6.88 (s, 1 arom. H); 7.27 (d, J ¼ 8.0, 2 arom. H); 7.36 (d, J ¼ 8.0, 2 arom.
H); 9.49 (s, NH). 13C-NMR (CDCl3): 13.67 (MeCH2); 19.15 (MeCH2); 21.22 (2 Me); 33.29 (ArCH2);
58.14 (CH2C ꢀ ); 64.70 (CH2OH); 72.78 (CH2N); 84.36 (CH2C ꢀ C); 86.32 (CH2C ꢀ C); 116.87 (C(5));
121.20; 124.99; 126.70; 128.99; 131.93; 134.79; 138.89; 141.63 (arom. C); 149.30 (C(2)); 152.00 (C(6));
163.36 (C(4)). HR-MALDI-MS: 441.1772 ([M þ Na]þ, C25H26N2NaO4þ ; calc. 441.1785). Anal. calc. for
C25H26N2O4 · 2.0 H2O (454.53): C 66.06, H 5.77, N 6.16; found: C 66.34, H 5.72, N 6.06.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-[({3-[3-(hydroxymethyl)phenyl]prop-2-ynyl}oxy)methyl]uracil
(27a). Yield: 129 mg (80%). White foam. Rf (3% MeOH/CHCl3) 0.23. 1H-NMR (CDCl3): 0.85 (t, J ¼ 7.4,
MeCH2); 2.25 (s, 2 Me); 2.37 (q, J ¼ 7.4, MeCH2); 4.10 (s, ArCH2); 4.52 (s, CH2C ꢀ ); 4.65 (s, CH2OH);
5.30 (s, CH2N); 6.69 (s, 2 arom. H); 6.88 (s, 1 arom. H); 7.22 – 7.26 (m, 3 arom. H); 7.41 (s, 1 arom. H); 9.83
(s, NH). 13C-NMR (CDCl3): 13.43 (MeCH2); 19.03 (MeCH2); 21.24 (2 Me); 33.23 (ArCH2); 58.42
(CH2C ꢀ ); 64.53 (CH2OH); 73.19 (CH2N); 84.68 (CH2C ꢀ C); 86.26 (CH2C ꢀ C); 116.94 (C(5)); 122.18;
125.01; 127.24; 128.35; 129.00; 130.40; 130.56; 134.81; 138.91; 141.58 (arom. C); 149.67 (C(2)); 152.22
(C(6)); 163.78 (C(4)). HR-MALDI-MS: 441.1774 ([M þ Na]þ, C25H26N2NaO4þ ; calc. 441.1785). Anal.
calc. for C25H26N2O4 · 1.5 H2O (445.52): C 67.40, H 5.88, N 6.29; found: C 67.94, H 5.83, N 6.28.
General Procedure for the Preparation of Aldehydes 28a and 29a. Alcohol 26a (86 mg, 0.2 mmol) was
dissolved in AcOEt (10 ml), and IBX (167 mg, 0.6 mmol) was added. The resulting suspension was
immersed in an oil-bath set to 808 and stirred vigorously until TLC showed no more starting material (3 –
4 h). After cooling, the mixture was filtered through a medium glass frit. The volatiles were removed
under reduced pressure, and the residue purified by CC (SiO2; CHCl3).
4-(3-{[6-(3,5-Dimethylbenzyl)-5-ethyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl]methoxy}prop-1-
ynyl)benzaldehyde (28a). Yield: 84 mg (97%). Yellow foam. Rf (3% MeOH/CHCl3) 0.39. 1H-NMR
(CDCl3): 1.01 (t, J ¼ 7.4, MeCH2); 2.25 (s, 2 Me); 2.45 (q, J ¼ 7.4, MeCH2); 4.10 (s, ArCH2); 4.54 (s,
CH2C ꢀ ); 5.28 (s, CH2N); 6.70 (s, 2 arom. H); 6.89 (s, 1 arom. H); 7.53 (d, J ¼ 7.5, 2 arom. H); 7.80 (d, J ¼
7.5, 2 arom. H); 9.59 (s, NH); 9.98 (s, CHO). 13C-NMR (CDCl3): 13.65 (MeCH2); 19.14 (MeCH2); 21.24
(2 Me); 33.32 (ArCH2); 57.96 (CH2C ꢀ ); 72.75 (CH2N); 85.51 (CH2C ꢀ C); 88.35 (CH2C ꢀ C); 116.93
(C(5)); 124.96; 129.06; 129.48; 132.30; 134.65; 135.74; 138.96 (arom. C); 149.38 (C(6)); 151.98 (C(2));
163.47 (C(4)); 191.34 (CHO). HR-MALDI-MS: 453.1785 ([M þ Na]þ, C26H26N2NaO4þ ; calc. 453.1785).
3-(3-{[6-(3,5-Dimethylbenzyl)-5-ethyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl]methoxy}prop-1-
ynyl)benzaldehyde (29a). Compound 29a was prepared in analogous way as 28a starting with 27a
(103 mg, 0.24 mmol) and IBX (200 mg, 0.71 mmol) in AcOEt (10 ml). Yield: 100 mg (98%). Yellow
1
foam. Rf (3% MeOH/CHCl3) 0.39. H-NMR (CDCl3): 1.00 (t, J ¼ 7.4, MeCH2); 2.25 (s, 2 Me); 2.44 (q,
J ¼ 7.4, MeCH2); 4.10 (s, ArCH2); 4.53 (s, CH2C ꢀ ); 5.28 (s, CH2N); 6.70 (s, 2 arom. H); 6.89 (s, 1 arom.
H); 7.45 – 7.50 (m, 1 arom. H); 7.61 – 7.65 (m, 1 arom. H); 7.81 – 7.84 (m, 1 arom. H); 7.89 – 7.90 (m, 1 arom.
H); 9.60 (s, NH); 9.97 (s, CHO). 13C-NMR (CDCl3): 13.66 (MeCH2); 19.14 (MeCH2); 21.23 (2 Me);
33.32 (ArCH2); 57.93 (CH2C ꢀ ); 72.71(CH2N); 84.00 (CH2C ꢀ C); 86.00 (CH2C ꢀ C); 116.93 (C(5));
124.97; 129.05; 129.10; 129.28; 133.20; 134.69; 136.40; 137.28; 138.94 (arom.); 149.41 (C(6)); 152.02
(C(2)); 163.47 (C(4)); 191.38 (CHO). HR-MALDI-MS: 453.1773 ([M þ Na]þ, C26H26N2NaO4þ ; calc.
453.1785).
Synthesis of 1-({[2,3-Bis(phenylsulfanyl)prop-2-enyl]oxy}methyl)-6-(3,5-dimethylbenzyl)-5-ethylur-
acil (30). Ph2S2 (72 mg, 0.33 mmol) and PPh3 (13 mg, 0.05 mmol) were stirred in a 5-ml round-bottomed
flask at 1008 for 2 min until homogenous yellow melt was obtained. Pd(Ph3)4 (5.8 mg, 0.005 mmol) was
added to the flask, and the mixture was kept stirring for 1 min. Finally, 1a (163 mg, 0.5 mmol) was added,
and the mixture was kept at 1008 for 2 h under stirring. After cooling, the residue was dissolved in CHCl3
and loaded on a SiO2 column. Elution with CHCl3 gave pure 30 (156 mg, 88%). White foam. 1H-NMR
(CDCl3): 1.08 (t, J ¼ 7.4, MeCH2); 2.26 (s, 2 Me); 2.44 (q, J ¼ 7.4, MeCH2); 4.02 (s, ArCH2); 4.16 (s,
CH2O); 5.05 (s, CH2N); 6.64 (s, 2 arom. H); 6.87 (s, 1 arom. H); 7.04 (s, CH¼); 7.21 – 7.45 (m, 10 arom.
H); 9.07 (s, NH). 13C-NMR (CDCl3): 13.75 (MeCH2); 19.18 (MeCH2); 21.27 (2 Me); 33.21 (ArCH2);
71.76 (CH2N); 72.32 (CH2O); 116.77 (C(5)); 125.00; 126.14; 126.90; 127.51; 128.94; 129.07; 129.22;
129.87; 130.44; 132.89; 133.19; 134.52; 134.85; 137.94; 138.84 (arom. C); 149.32 (C(2)); 151.73 (C(6));
163.12 (C(4)). HR-MALDI-MS: 567.1730 ([M þ Na]þ, C31H32N2NaO3Sþ2 ; calc. 567.1747). Anal. calc. for
C31H32N2O3S2 · 0.25 H2O (549.24): C 67.79, H 5.96, N 5.10; found: C 67.46, H 5.94, N 5.03.

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Helvetica Chimica Acta – Vol. 92 (2009)
General Procedure for the ꢂClickꢀ Chemistry on 1a. A small flask (5 ml) was charged with 1a (163 mg,
0.5 mmol), the appropriate halo compound (0.5 mmol), (S)-proline (12 mg, 0.1 mmol), Na2CO3 (12 mg,
0.1 mmol), NaN3 (42 mg, 0.65 mmol), and CuSO4 · 5 H2O (13 mg, 0.025 mmol). To the mixture was
added DMSO/H2O 9 :1 (2 ml), followed by freshly prepared 1m aq. soln. of sodium ascorbate (0.1 ml, ca.
25 mg, 0.05 mmol). The mixture was heated at 658 overnight with vigorous stirring. After cooling to r.t.,
the precipitate was filtered (washing with dil. NH3 and H2O).
6-(3,5-Dimethylbenzyl)-5-ethyl-1-({[1-(pyridin-3-yl)-1H-[1,2,3]triazol-4-yl]methoxy}methyl)uracil
(31). Yield: 205 mg (92%). White solid. Rf (5% MeOH/CHCl3) 0.27. M.p. 197 – 1988. 1H-NMR (CDCl3):
1.09 (t, J ¼ 7.4, MeCH2); 2.26 (s, 2 Me); 2.48 (q, J ¼ 7.4, MeCH2); 4.10 (s, ArCH2); 4.86 (s, CH2O); 5.19 (s,
CH2N); 6.69 (s, 2 arom. H); 6.88 (s, 1 arom. H); 7.47 – 7.52 (m, 1 H of pyridinyl); 8.20 (d, J ¼ 9.0, 1 H of
pyridinyl); 8.35 (s, HꢂC(5) of triazolyl); 8.70 (br. s, 1 H of pyridinyl); 9.09 (br. s, 1 H of pyridinyl); 9.78
(s, NH). 13C-NMR (CDCl3): 13.71 (MeCH2); 19.27 (MeCH2); 21.26 (2 Me); 33.40 (ArCH2); 61.90
(CH2O); 71.79 (CH2N); 117.29 (C(5)); 121.84; 124.28; 125.00; 128.11; 129.05; 133.62; 134.66; 138.95;
141.65; 144.68 (arom. C); 149.20 (C(2)); 149.89 (arom. C); 152.67 (C(6)); 163.07 (C(4)). HR-MALDI-
MS: 469.1938 ([M þ Na]þ, C24H26N6NaO3þ ; calc. 469.1959). Anal. calc. for C24H26N6O3 · 0.25 H2O
(451.02): C 63.92, H 5.81, N 18.63; found: C 63.72, H 5.73, N 18.50.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-({[1-prop-2-enyl)-1H-[1,2,3]triazol-4-yl]methoxy}methyl)uracil
(32). Yield: 170 mg (41%). White solid. Rf (3% MeOH/CHCl3) 0.27. M.p. 167 – 1698. 1H-NMR (CDCl3):
1.08 (t, J ¼ 7.5, MeCH2); 2.26 (s, 2 Me); 2.47 (q, J ¼ 7.5, MeCH2); 4.07 (s, ArCH2); 4.75 (s, CH2O); 4.98 (d,
J ¼ 6.3, NCH2CH¼); 5.15 (s, CH2N); 5.29 – 5.36 (m, ¼CH2); 5.96 – 6.07 (m, NCH2CH¼); 6.67 (s, 2 arom.
H); 6.88 (s, 1 arom. H); 7.73 (s, HꢂC(5) of triazolyl); 9.51 (s, NH). 13C-NMR ((D6)DMSO): 13.50
(MeCH2); 18.51 (MeCH2); 20.77 (2 Me); 32.71 (ArCH2); 51.53 (NCH2CH¼); 61.54 (CH2O); 71.82
(CH2N); 115.40 (C(5)); 118.71 (NCH2CH¼); 123.84; 124.80; 128.21 (arom. C); 132.55
(NCH2CH¼CH2); 135.72; 137.90; 143.20 (arom. C); 148.16 (C(2)); 151.45 (C(6)); 162.92 (C(4)). HR-
MALDI-MS: 432.1988 ([M þ Na]þ, C22H27N5NaO3þ ; calc. 432.2006).
1-{[(1-Benzyl-1H-[1,2,3]triazol-4-yl)methoxy]methyl}-6-(3,5-dimethylbenzyl)-5-ethyluracil (33).
Yield: 147 mg (64%). White solid. Rf (3% MeOH/CHCl3) 0.30. M.p. 137 – 1398. 1H-NMR (CDCl3):
1.08 (t, J ¼ 7.5, MeCH2); 2.26 (s, 2 Me); 2.46 (q, J ¼ 7.5, MeCH2); 4.06 (s, ArCH2); 4.71 (s, CH2O); 5.11 (s,
CH2N); 5.54 (s, NCH2Ph); 6.66 (s, 2 arom. H); 6.87 (s, 1 arom. H); 7.33 (s, 5 arom. H); 7.72 (s, HꢂC(5) of
triazolyl); 9.74 (s, NH). 13C-NMR (CDCl3): 13.69 (MeCH2); 19.20 (MeCH2); 21.24 (2 Me); 33.31
(ArCH2); 54.03 (NCH2Ph); 62.06 (CH2O); 71.83 (CH2N); 117.07 (C(5)); 123.55; 124.98; 128.13; 128.65;
128.96; 129.01; 134.70; 138.85; 143.75 (arom. C); 149.24 (C(2)); 152.48 (C(6)); 163.14 (C(6)). HR-
MALDI-MS: 482.2152 ([M þ Na]þ, C26H29N5 NaO3þ ; calc. 482.2163).
1-({[1-(2,6-Difluorobenzyl)-1H-[1,2,3]triazol-4-yl]methoxy}methyl)-6-(3,5-dimethylbenzyl)-5-ethyl-
uracil (34). Yield: 78 mg (12%). White solid. Rf (3% MeOH/CHCl3) 0.27. M.p. 161 – 1638. 1H-NMR
(CDCl3): 1.07 (t, J ¼ 7.5, MeCH2); 2.26 (s, 2 Me); 2.46 (q, J ¼ 7.5, MeCH2); 4.05 (s, ArCH2); 4.72 (s,
CH2O); 5.14 (s, CH2N); 5.63 (s, NCH2Ar); 6.66 (s, 2 arom. H); 6.87 (s, 1 arom. H); 6.93 – 6.98 (m, 2 arom.
H); 7.33 – 7.38 (m, 1 arom. H); 7.73 (s, HꢂC(5) of triazolyl); 9.43 (s, NH). 13C-NMR (CDCl3): 13.67
(MeCH2); 19.18 (MeCH2); 21.23 (2 Me); 33.29 (ArCH2); 41.27 (t, J ¼ 3.9, CH2N); 62.28 (CH2O); 72.20
(CH2N); 110.99 (t, J ¼ 18.7); 111.79 (dd, J ¼ 17.5, 7.2); 116.93 (C(5)); 123.33, 124.98, 128.94, 131.36 (t, J ¼
10.3); 134.72; 138.85; 143.81; 149.24 (C(2)); 152.15 (C(6)); 161.33 (dd, J ¼ 250.0, 6.9); 163.15 (C(4)). HR-
MALDI-MS: 518.1956 ([M þ Na]þ, C26H27F2N5NaO3þ ; calc. 518.1974).
General Procedure for the Bromination of 2a and 2b to give 35a and 35b, Resp. To a soln. of 2a or 2b
(288 mg, 0.74 mmol) in dry CH2Cl2 (25 ml) was dropwise added a 0.78m soln. of Br2 in CH2Cl2
(0.78 mmol, 1 ml) with stirring at 08. After the addition was completed, stirring was continued for
30 min at 08. The solvent was removed under reduced pressure, and the residue was chromatographed on
SiO2 (CHCl3).
1-{[(2,3-Dibromo-3-phenylprop-2-enyl)oxy]methyl}-6-(3,5-dimethylbenzyl)-5-ethyluracil (35a).
1
Yield: 272 mg (72%). White foam. Rf (CHCl3) 0.16. H-NMR (CDCl3): 1.07 (t, J ¼ 7.4, MeCH2); 2.29
(s, 2 Me); 2.49 (q, J ¼ 7.4, MeCH2); 4.16 (s, ArCH2); 4.74 (s, CH2O); 5.27 (s, CH2N); 6.74 (s, 2 arom. H);
6.91 (s, 1 arom. H); 7.32 – 7.39 (m, 5 arom. H); 9.54 (s, NH). 13C-NMR (CDCl3): 13.81 (MeCH2); 19.15
(MeCH2); 21.29 (2 Me); 33.33 (ArCH2); 72.78 (CH2N); 73.73 (CH2O); 116.97 (C(5)); 118.07
(CH2CBr¼CBr); 120.99 (CH2CBr¼CBr); 125.06; 128.30; 128.72; 129.01; 129.09; 134.80; 138.92;

Helvetica Chimica Acta – Vol. 92 (2009)
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139.85 (arom. C); 149.33 (C(2)); 151.90 (C(6)); 163.30 (C(4)). HR-MALDI-MS: 583.0191 ([M þ Na]þ,
C25H26Br2N2NaOþ3 ; calc. 583.0202). Anal. calc. for C25H26Br2N2O3 · 0.20 H2O (565.92): C 53.01, H 4.62, N
4.95; found: C 53.07, H 4.55, N 4.73.
1-{[(2,3-Dibromo-3-phenylprop-2-enyl)oxy]methyl}-5-ethyl-6-(2-methylbenzyl)uracil (35b). Yield:
262 mg (65%). White foam. Rf (CHCl3) 0.17. 1H-NMR (CDCl3): 1.07 (t, J ¼ 7.4, MeCH2); 2.29 (s, 2 Me);
2.49 (q, J ¼ 7.4, MeCH2); 4.16 (s, ArCH2); 4.74 (s, CH2O); 5.27 (s, CH2N); 6.74 (s, 2 arom. H); 6.91 (s, 1
arom. H); 7.32 – 7.39 (m, 5 arom. H); 9.54 (s, NH). 13C-NMR (CDCl3): 13.68 (MeCH2); 19.11 (MeCH2);
19.76 (2-MeC6H4); 30.99 (ArCH2); 73.15 (CH2N); 73.74 (CH2O); 117.24 (C(5)); 118.13 (CH2CBr¼CBr);
120.81 (CH2CBr¼CBr); 125.63, 126.78, 127.33, 128.52, 129.09, 129.48, 130.68, 133.35, 136.16, 139.86
(arom. C); 149.49 (C(2)); 151.77 (C(6)); 163.02 (C(4)). HR-MALDI-MS: 569.0057 ([M þ Na]þ,
C24H24Br2N2NaOþ3 ; calc. 569.0046). Anal. calc. for C24H24Br2N2O3 · 0.25 H2O (552.79): C 52.15, H 4.47, N
5.07; found: C 52.19, H 4.42, N 4.74.
6-(3,5-Dimethylbenzyl)-5-ethyl-1-({[4-(3-hydroxyprop-1-ynyl)benzyl]oxy}methyl)uracil (37). Com-
pound 37 was prepared from 36 [21] and propargyl alcohol according to the general procedure for
Sonogashira coupling. Yield: 140 mg (95%). Beige foam. Rf (3% MeOH/CHCl3) 0.26. 1H-NMR
(CDCl3): 1.06 (t, J ¼ 7.4, MeCH2); 2.27 (s, 2 Me); 2.45 (q, J ¼ 7.4, MeCH2); 4.05 (s, ArCH2); 4.49 (s, ꢀ
CCH2OH); 4.63 (s, CH2O); 5.19 (s, CH2N); 6.64 (s, 2 arom. H); 6.88 (s, 1 arom. H); 7.25 (d, J ¼ 8.1, 2
arom. H); 7.39 (d, J ¼ 8.1, 2 arom. H); 9.39 (s, NH). 13C-NMR (CDCl3): 13.74 (MeCH2); 19.14 (MeCH2);
21.25 (2 Me); 33.25 (ArCH2); 51.53 (C ꢀ CCH2OH); 71.27 (CH2O); 72.82 (CH2N); 85.24 (C ꢀ
CCH2OH); 87.64 (C ꢀ CCH2OH); 116.88 (C(5)); 122.20; 124.95; 127.59; 129.00; 131.73; 134.73; 137.71;
138.89 (arom. C); 149.15 (C(6)); 151.92 (C(2)); 163.26 (C(4)). HR-MALDI-MS: 455.1923 ([M þ Na]þ,
C26H28N2NaOþ4 ; calc. 455.1941). Anal. calc. for C26H28N2O4 · 0.75 H2O (446.03): C 69.95, H 6.28, N 6.28;
found: C 69.78, H 6.36, N 6.19.
3-[4-({[6-(3,5-Dimethylbenzyl)-5-ethyl-2,4-dioxo-3,4-dihydro-2H-pyrimidin-1-yl]methoxy}methyl)-
phenyl]propynal (38). Compound 38 was prepared from 37 according to the procedure for compounds
28a and 29a. Yield: 59 mg (76%). Yellow foam. Rf (3% MeOH/CHCl3) 0.39. 1H-NMR (CDCl3): 1.06 (t,
J ¼ 7.4, MeCH2); 2.27 (s, 2 Me); 2.45 (q, J ¼ 7.4, MeCH2); 4.07 (s, ArCH2); 4.69 (s, CH2O); 5.22 (s, CH2N);
6.67 (s, 2 arom. H); 6.89 (s, 1 arom. H); 7.35 (d, J ¼ 8.4, 2 arom. H); 7.39 (d, J ¼ 8.4, 2 arom. H); 9.32 (s,
NH); 9.41 (s, CHO). 13C-NMR (CDCl3): 13.72 (MeCH2); 19.15 (MeCH2); 21.26 (2 Me); 33.30 (ArCH2);
70.95 (CH2O); 72.88 (CH2N); 88.54 (C ꢀ CCHO); 94.73 (C ꢀ CCHO); 117.00 (C(5)); 118.83; 124.92;
127.70; 129.05; 133.34; 134.64; 138.94; 141.00 (arom. C); 149.05 (C(6)); 151.91 (C(2)); 163.16 (C(4));
176.68 (CHO). HR-MALDI-MS: 453.1782 ([M þ Na]þ, C26H26N2NaO4þ ; calc. 453.1785).
Antiviral Assay Procedures. Compounds were solubilized in DMSO at 100 mm and then diluted in
culture medium.
Virus and Cells. MT-4, C8166, and H9/IIIB cells were grown at 378 in a 5% CO2 atmosphere in RPMI
1640 medium supplemented with 10% fetal calf serum (FCS), 100 IU/ml penicillin G, and 100 mg/ml
streptomycin. Cell cultures were checked periodically for the absence of mycoplasma contamination with
a MycoTect Kit (Gibco). Human immunodeficiency viruses type 1 (HIV-1, IIIB strain) was obtained
from supernatants of persistently infected H9/IIIB cells. The HIV-1 stock solns. had titers of 4.5 ꢃ 106
50% cell culture infectious dose (CCID50)/ml. The Y181C mutant (NIH N119) was derived from an
AZT-sensitive clinical isolate passaged initially in CEM and then in MT-4 cells in the presence of
nevirapine (10 mm). The double mutant K103N þ Y181C (NIH A17) was derived from the IIIB strain
passaged in H9 cells in the presence of BI-RG 587 (1 mm). The triple mutant K103R þ V179D þ P225 H
(EFVR) was derived from an IIIB strain passaged in MT-4 cells in the presence of efavirenz (up to 2 mm).
N119, A17, and EFVR stock solns. had titers of 1.2 ꢃ 108, 2.1 ꢃ 107, and 4.0 ꢃ 107 CCID50/ml, resp.
HIV Titration. Titration of HIV was performed in C8166 cells by the standard limiting dilution
method (dilution 1:2, four replica wells per dilution) in 96-well plates. The infectious virus titer was
determined by light microscope scoring of syncytia after 4 d of incubation. Virus titers were expressed as
CCID50/ml.
Anti-HIV Assays. The activity of test compounds against multiplication of HIV-1 wild type IIIB,
N119, A17, and EFVR in acutely infected cells was based on inhibition of virus-induced cytopathicity in
MT-4 cells. Briefly, an amount of 50 ml of culture medium containing 1 ꢃ 104 cells was added to each well
of flat-bottom microtiter trays containing 50 ml of culture medium with or without various concentrations

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Helvetica Chimica Acta – Vol. 92 (2009)
of test compounds. Then, an amount of 20 ml of HIV-1 suspensions (containing the appropriate amount
of CCID50 to cause complete cytopathicity at day 4 was added. After incubation at 378, cell viability was
determined by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium hydrobromide (MTT)
method [60]. The cytotoxicity of test compounds was evaluated in parallel with their antiviral activity
and was based on the viability of mock-infected cells, as monitored by the MTT method.
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Received February 3, 2009