Synthesis of Tetrasubstituted Thiophenes via Direct Metalation

Article abstract of DOI:10.1021/acs.joc.9b02803

Thiophene moiety can be derivatized by various synthetic procedures. The most convenient method seems to be derivatization via direct metalation, but synthesis of polysubstituted thiophenes bearing reactive groups is difficult because of high reactivity of organometallic reagents. This work reports the preparation of complex heterocyclic compounds using direct metalation of thiophenes with various reagents (Knochel-Hauser bases, LDA) as an efficient synthetic tool.

Full text of DOI:10.1021/acs.joc.9b02803

Subscriber access provided by Chalmers Library
Article
Synthesis of Tetrasubstituted Thiophenes via Direct Metalation
Jan Skácel, Martin Dracinsky, and Zlatko Janeba
J. Org. Chem., Just Accepted Manuscript • DOI: 10.1021/acs.joc.9b02803 • Publication Date (Web): 20 Dec 2019
Downloaded from pubs.acs.org on December 22, 2019
Just Accepted
“Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted
online prior to technical editing, formatting for publication and author proofing. The American Chemical
Society provides “Just Accepted” as a service to the research community to expedite the dissemination
of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in
full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully
peer reviewed, but should not be considered the official version of record. They are citable by the
Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore,
the “Just Accepted” Web site may not include all articles that will be published in the journal. After
a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web
site and published as an ASAP article. Note that technical editing may introduce minor changes
to the manuscript text and/or graphics which could affect content, and all legal disclaimers and
ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or
consequences arising from the use of information contained in these “Just Accepted” manuscripts.
is published by the American Chemical Society. 1155 Sixteenth Street N.W.,
Washington, DC 20036
Published by American Chemical Society. Copyright © American Chemical Society.
However, no copyright claim is made to original U.S. Government works, or works
produced by employees of any Commonwealth realm Crown government in the
course of their duties.

Page 1 of 27
The Journal of Organic Chemistry
1
2
3
4
5
Synthesis of Tetrasubstituted Thiophenes via Direct Metalation
6
7
8
9
Jan Skácel, Martin Dračínský, Zlatko Janeba*
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Institute of Organic Chemistry and Biochemistry of the Czech Academy of Sciences,
Flemingovo nám. 2, CZ-16610 Prague 6, Czech Republic
E-mail: janeba@uochb.cas.cz
Abstract
OBn
Boc
4 steps
up to 44 % overall yield
N
N
S
N
P(O)(OR)2
n
S
Br
Br
EtOOC
S
Z = COOEt, P(O)(OR)2
n = 1, 2; R = Et, i-Pr
Z
Thiophene moiety can be derivatized by various synthetic procedures. The most convenient
method seems to be derivatization via direct metalation but synthesis of polysubstituted
thiophenes bearing reactive groups is difficult due to high reactivity of organometallic reagents.
This work reports the preparation of complex heterocyclic compounds using direct metalation
of thiophenes with various reagents (Knochel-Hauser bases, LDA) as an efficient synthetic tool.
1
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 2 of 27
1
2
3
4
5
6
INTRODUCTION
In our research group, we focus on a study of modified nucleotide analogues for medical use in
order to treat various viral1 or parasitic2–8 infections, among others. Recently, we were interested
in structures, which would mimic geometry and charge distribution of transition states involved
in the enzymatic transformation of 6-oxopurines and phosphoribosyl pyrophosphate, as natural
substrates, by phosphoribosyl transferases (Figure 1). Hypoxanthine-guanine-(xanthine)
phosphoribosyltransferase (HG(X)PRT), for example, represents a recognized target for
development of potential anti-malarial chemotherapeutics.
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
O
N
O
OBn
N
Boc
H
N
N
N
N
NH
NH
O
-O P O
O-
O
N
O
N
O
S
S
P
-O
(EtO)2(O)P
*
O-
O
O
COOEt
S
O-
S
OH OH
P
O-
O-
O
O
O
P
P(O)(OEt)2
Target molecule
P
O
O
O
Artificial mimic
HG(X)PRT transition state
Figure 1. Comparison of geometry and charge distribution between the HG(X)PRT transition
state and our target molecule
We have attempted to design artificial nucleotide mimics based on a thiophene moiety
(Figure 1), which could be exploited as a tool to search for novel medicinal targets and new
therapeutic opportunities. The Reaxys database contains approx. 900k compounds derived from
the thiophene moiety, but only approx. 21k molecules contain the tetrasubstituted thiophene
core and, moreover, mostly in a symmetrical fashion. To our surprise, no structures similar to
our target molecule (Figure 1) were present in available databases, although thiophene moiety
is commonly part of agrochemicals, pharmaceuticals and electronics based on organic
molecules.9–12 Therefore, we have developed a new synthetic strategy for a fully
regioselectivelly controlled synthesis of asymmetrically substituted thiophenes.
Thiophene derivatives are usually synthesized by aromatic electrophilic substitution13 or by
heterocyclization methods,14 but these approaches require additional transformations, which
increase number of steps in the synthesis. In addition, standard transformations often require
energetic conditions not compatible with sensitive and/or reactive functional groups. Thus, we
employed a straightforward introduction of functional groups by direct metalation combined
2
ACS Paragon Plus Environment

Page 3 of 27
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
with subsequent reactions of intermediates with corresponding electrophiles.15 As the key
reagents, Knochel-Hauser bases, like tetramethylpiperidinylmagnesium chloride lithium
chloride complex (TMPMgCl‧LiCl),16 were used. Such mixed amide base exhibits remarkable
functional group tolerance and allows straightforward synthesis of tetrasubstituted
heterocycles.17,18 However, the complexity of final molecules was quite limited. In this work,
we explored applications of amide bases for selective, late-stage deprotonation of complex
molecules.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
RESULTS AND DISCUSSION
Since no similarly polysubstituted structures (Figure 1) have been reported, the questions
regarding to stability of the target molecules arose. We needed a proof-of-concept synthesis.
First, amino thiophene (1, Scheme 1) was prepared in a 75% yield by the Gewald
condensation.19 Compound 1 was then converted into 2-bromothiophene (2)20 by diazotation
and subsequent reaction with copper (II) bromide. The diazotation step proceeded in variable
yields (20% – 50%) and turned-out to be difficult to optimize. The bromine substituent was
transformed catalytically to diethyl phosphonate derivative 3 (75%).21 Bromination of substrate
3 under various reaction conditions led to completely different products. Halogenation of 3 with
NBS in acetonitrile at 80 °C without radical initiator led exclusively to the formation of
bromomethyl derivative 4. When catalytic amount of acetic or trifluoroacetic acid was used,
the reaction afforded a mixture of products (we assume that the diethyl phosphonate moiety
was converted into free phosphonic acid, which acted as a leaving group and got substituted by
bromine). On the other hand, bromination in an amber flask using bromine in acetic acid at
room temperature led to a formation of the desired 4-bromothiophene 5 in a quantitative yield.
In order to prepare boronic ester 6, derivative 5 was subjected to Pd-catalyzed borylation with
Pd(Pt-Bu3)2 and pinacolborane in the presence of triethylamine. Nevertheless, boronate 6 could
not be isolated since (hetero)aromatic boronic acids/esters bearing electron withdrawing groups
are usually prone to protodeboronation.22,23 Thus, boronate 6 was used in the next reaction
without isolation.
3
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 4 of 27
1
2
3
4
5
6
7
i-AmONO,
CuBr2
EtOOC
O
OEt
CN
H
O
EtOOC
S
S
S8, Et3N
+
H2N
Br
_
ACN,
-10 °C  rt
EtOH, DMF,
5 h, rt  60 °C
S8
2
, 20% 50%
8
1
, 75%
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Br
HP(O)(OEt)2, Pd2(dba)3,
Xantphos, Et3N
EtOOC
(EtO)2(O)P
EtOOC
S
NBS, ACN
14 h, 80 °C
S
(EtO)2(O)P
dioxane, 2 h, 80 °C
4
, 80%
3
, 81%
AcOH
rt
Br2,
dark,
h,
14
OBn
Boc
N
N
Br
HB(Pin), Et3N,
9
, XPhos G2,
K3PO4
O
N
O
Pd(t-Bu3P)2
B
EtOOC
S
THF, 2 h,
40 °C
THF/H2O,
1 h, 40 °C
EtOOC
(EtO)2(O)P
S
EtOOC
(EtO)2(O)P
S
(EtO)2(O)P
5
, 96%
6
7
, 50%
Scheme 1. Synthesis of target tetrasubstituted thiophene 7.
Bromo derivative 9 (Scheme 2) was chosen as a reaction partner for compound 6 (Scheme 1)
in the subsequent Suzuki coupling. Compound 9 was synthesized from 6-chloro-9-deazapurine
(8) by nucleophilic substitution with benzyl alcoholate, followed by electrophilic bromination
with NBS and introduction of Boc group to 7-NH position. The pyrrole ring in 9-deazapurines
is relatively electron rich, which is generally problematic for Pd-catalyzed couplings. Thus, the
Boc group was used here to deplete part of electron density since the carbamate group is more
electron-withdrawing group compared to amide- or sulfonamide-containing protecting
groups.24
OBn
Cl
Boc
H
(i) BnOH, Na (96%)
N
N
N
N
(ii) NBS, THF, 1 h, rt,
(iii) Boc2O, DMAP, ACN
2 h, 40 °C (81%)
N
N
Br
, 78%
8
9
Scheme 2. Synthesis of suitably protected 9-bromo-9-deazapurine 9.
The optimized procedure for catalytic borylation of bromothiophene 5, followed by filtration
of the reaction mixture through celite and evaporation afforded crude 6. Compound 6 was used
4
ACS Paragon Plus Environment

Page 5 of 27
The Journal of Organic Chemistry
1
2
3
4
5
6
directly in Suzuki coupling with 9-deazapurine 9 to offer target product 7 (Scheme 1) in a 50%
yield (from 5).
7
8
9
The direct metalation was chosen for the synthesis of desired tetrasubstituted thiophenes.
Because of presence of reactive or sensitive functional groups, the selected metalating reagent
should react under mild reaction conditions. Such requirements were completely implemented
by use of TMPMgCl‧LiCl. This base was used previously for the synthesis of polysubstituted
aromatic compounds.25–28
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
First, we selected metalation of 2-benzoylthiothiophene 11 (Scheme 3) which was prepared
quantitatively using the Ullmann coupling of 2-iodothiophene (10) and thiobenzoic acid.29
Then, ethyl carboxylate moiety was introduced in a high yield by the reaction of the metalated
intermediate and ethyl cyanoformate, to give compound 12. Nevertheless, the second
metalation on 12 only led to a cleavage of the benzoyl group, resulting in a complex product
mixture (including traces of 13). These results suggested that a successful synthetic strategy
would require using 3-bromo substituted thiophenes as the starting material.
PhCOSH, Et3N, CuI,
1,10-phenantoline
(i) TMPMgCl.LiCl, THF,
30 min, -25 °C
SBz
I
S
S
toluene,
1 h, 100 °C
(ii) CNCOOEt, THF,
30 min, -25 °C  rt
11
, 98%
10
SBz
SBz
(i) TMPMgCl.LiCl, THF,
30 min, -25 °C
EtOOC
EtOOC
S
S
(ii) CNCOOEt, THF,
30 min, -25 °C  rt
13
traces (UPLC)
EtOOC
12
, 99%
Scheme 3. An attempt to prepare compound 13 using direct metalation.
Commercially available 3,4-dibromothiophene (14, Scheme 4) was used for the halogen-metal
exchange preparation of the Grignard reagent and its subsequent reaction with ethyl
chloroformate to obtain ethyl 4-bromothiophene-3-carboxylate (15) in a 82% yield. This
substitution activated C2 position of thiophene 15 towards Knochel-Hauser base.
Deprotonation of 15 followed by the reaction with ethyl cyanoformate gave trisubstituted
thiophene 16 in an excellent yield. The last substituent was introduced by deprotonation of 16
and by subsequent reaction of organometallic C5-intermediate with sulfur. In-situ alkylation of
formed thiolate with diisopropyl triflyloxymethylphosphonate afforded the desired
tetrasubstituted thiophene 17 in a 58% overall yield (4 reaction steps).
5
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 6 of 27
1
2
3
4
(i) i-PrMgBr, THF,
(i) TMPMgCl.LiCl, THF,
S
S
5 h, 0 °C
30 min, -20 °C
5
6
7
(ii) ClCOOEt,
14 h, rt
(ii) CNCOOEt, THF,
30 min, -20  0 °C,
Br
Br
Br
COOEt
14
15
, 82%
8
(i-PrO)2(O)P
9
S
(i) TMPMgCl.LiCl, THF,
30 min., -20 °C
S
COOEt
S
COOEt
COOEt
TfOCH2P(O)(Oi-Pr)2
THF, 10 min, 0 °C
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(ii) S8, THF, 30 min,
Br
17
COOEt
, 98%
14
Br
-20  0 °C
16
, 72%
58% from
Scheme 4. Synthesis of tetrasubstituted thiophene 17 using direct metalation.
In contrast, the reaction of metalated intermediate of compound 15 with diethyl
chlorophosphate afforded the desired diethyl thiophene-2-phosphonate 18 (Scheme 5) in a low
yield (< 10% by UPLC). Therefore, halogenation of the C2 position followed by the
Pd-catalyzed introduction of diethyl phosphonate group was studied and optimized.
2-Iodothiophene 19 and 2-bromothiophene 20 (Scheme 5) was prepared in high yields by the
reaction of metalated 15 with iodine or 1,2-dibromo-1,1,2,2-tetrachloroethane, respectively.
Subsequent coupling of 2-halothiophenes with diethyl phosphite was much more robust in the
case of 2-bromo derivative 20 (85% of 18) than in the case of 2-iodo derivative 19 (14–60% of
18). The coupling with 2-iodo derivative 19 was extremely sensitive towards oxygen and/or
water residues and it suffered from major dehalogenation of iodine in starting material as well
as from cleavage of ethyl ester groups from the diethyl phosphonate moiety in product 18. The
yields were variable and such a method was not applicable for multigram scale-up. In contrast,
the procedure using 2-bromo derivative 20 afforded desired compound 18 smoothly in high
yields even in multigram scale-ups.
6
ACS Paragon Plus Environment

Page 7 of 27
The Journal of Organic Chemistry
1
2
3
4
(i) TMPMgCl.LiCl, THF,
30 min, -20 °C
S
P(O)(OEt)2
5
6
7
(ii) ClP(O)(OEt)2, THF,
14 h, -20 °C  0 °C
Br
COOEt
18,
< 10% (UPLC)
8
9
S
S
HP(O)(OEt)2, Pd2(dba)3,
Xantphos, Et3N
(i) TMPMgCl.LiCl, THF,
30 min, -20 °C
I
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
dioxane, 2 h, 60 °C,
60%
(ii) I2, THF, 30 min,
COOEt
Br
19
COOEt
Br
-20 °C  0 °C
, 88%
15
HP(O)(OEt)2, Pd2(dba)3,
Xantphos, Et3N
S
S
(i) TMPMgCl.LiCl, THF,
30 min, -20 °C
P(O)(OEt)2
Br
dioxane, 2 h, 80 °C
(ii) (CCl2Br)2, THF,
30 min,
Br
18
COOEt
, 85%
Br
COOEt
, 81%
20
-20 °C  0 °C
Scheme 5. Synthetic approached to trisubstituted thiophene 18.
Compound 17, containing a single bromine substituent, was then used for the catalytic
borylation and subsequent Suzuki coupling. Although the borylation step worked with
reasonable conversion (UPLC), subsequent Suzuki coupling did not proceed and the reaction
resulted in deborylation. This may be caused by the steric hindrance of the borylated C3 position
and all attempts for the improvement failed. The same procedure worked perfectly with
compounds 16 and 18 (Scheme 6), lacking the heteroarylphosphonate moiety, and the reaction
provided desired products 24 and 26, respectively, in high yields.
OBn
Boc
S
Z
Y
C6
9
, Pd XPhos G2,
K3PO4
S
HB(Pin), Et3N,
Y
N
Z
N
Pd(t-Bu3P)2
C8
EtOOC
B O
C2
N
THF/H2O, 1 h,
40 °C
THF, 2 h,
40 °C
Y
EtOOC
Br
O
16
17
EtOOC
: Z = COOEt, Y = H
: Z = COOEt
Y = SCH2P(O)(Oi-Pr)2
: Z = P(O)(OEt)2, Y = H
S
21
22
: Z = COOEt, Y = H
: Z = COOEt
Y = SCH2P(O)(Oi-Pr)2
: Z = P(O)(OEt)2, Y = H
Z
24
25
: Z = COOEt, Y = H (82%)
: Z = COOEt
18
23
Y = SCH2P(O)(Oi-Pr)2 (< 3%)
: Z = P(O)(OEt)2, Y = H (76%)
26
Scheme 6. Synthesis of tetrasubstituted thiophenes bearing 9-deazapurine moiety.
Due to these complications, we decided to carry out metalation reactions with relatively
complex compounds 24 and 26. The 9-deazapurine moiety of both compounds contains acidic
protons at positions C2 and C8, nucleophilic sensitive positions C2 and C6, and relatively labile
Boc protecting group. Moreover, the thiophene moiety contains basic and nucleophilic sensitive
esters (carboxylate and phosphonate).
7
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 8 of 27
1
2
3
4
5
6
7
8
9
First, we decided to study the course of metalation with compound 24 using trapping with iodine
to obtain product 28 (Scheme 7). Despite the drawbacks described in the previous paragraph,
Knochel-Hauser base was able to selectively deprotonate only position C5 at the thiophene
moiety without formation of any observable side products. Nevertheless, yields of iodo
derivative 28 were below 20%, probably due to chelation of the magnesium ion by N3 nitrogen
in the metalated intermediate 27 (Scheme 7), which may interfere with the subsequent reaction
of the electrophile. The utilization of TMP2Mg, that keeps the bulky TMP ligand bound to the
magnesium ion, resulted only in a small improvement in the yields.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
OBn
OBn
OBn
Boc
Boc
Boc
TMPMgCl.LiCl
THF, -20 °C
I2
N
N
N
N
N
N
< 20%
N
I
N
N
COOEt
COOEt
COOEt
COOEt
COOEt
Mg
Cl
S
S
S
28
COOEt
24
27
(i) TMPLi, TMEDA, THF, 2 min, -78 °C,
(ii) I2, THF, 10 min, -78 °C  0 °C
72%
Scheme 7. A study of the metalation of compound 24 followed by trapping with iodine and
optimized synthesis of compound 28.
Logically, a replacement of the bivalent magnesium ion for the monovalent lithium ion was
studied next. Use of TMPLi at -78 °C provided higher, yet very variable, yields of lithiated
product (not shown). Further improvement was observed after a TMEDA addition but the yields
of lithiation were still quite variable. A careful study of the reaction course using UPLC
revealed that deprotonation of substrate 24 with TMPLi at -78 °C took only about 1 min and
the lithiated intermediate was stable for about 5 min, followed by its complete decomposition
within 40 min. Thus, addition of iodine 2 min after lithiation afforded compound 28 in a 72%
yield (Scheme 7).
Similarly, addition of sulfur within 1–2 min after lithiation provided desired intermediate 29
(Scheme 8) with high and reproducible conversion (UPLC). Subsequent in-situ alkylation of
29 with diisopropyl triflyloxymethylphosphonate delivered final compound 25 in a 68% yield
(4 isolated intermediates, 33% overall yield from 14). For the introduction of the
thioethylphosphonate moiety, it was necessary to treat intermediate 29 with hydrochloric acid
(to get intermediate 32) and then perform mild alkylation of the crude mixture with diethyl
2-bromoethylphosphonate in the presence of potassium carbonate in DMF. This procedure
8
ACS Paragon Plus Environment

Page 9 of 27
The Journal of Organic Chemistry
1
2
3
4
5
6
afforded final compound 34 (Scheme 8) in a 59% yield (5 isolated intermediates, 29% overall
yield from 14).
7
8
9
However, the above optimized reaction conditions did not work in the case of compound 26,
bearing the phosphonate moiety instead of the carboxylate at position C2 of thiophene. Simple
solution was to decrease the reaction temperature bellow -78 °C, down to -95 °C. This change
led to a dramatic increase of stability of the lithiated intermediate (UPLC), which was fully
stable for 30 min. In addition, it was possible to replace TMPLi/TMEDA for LDA as a
metalating agent, while magnesium amides were not reactive enough to deprotonate substrates
24 and 26 at temperatures below -40 °C. Moreover, larger excess of LDA resulted in multiple-
lithiated compounds (possible lithiation at positions C2, C8 of purine and at position C5 of
thiophene). The excess of TMPLi in the previous reaction conditions led to decomposition of
the starting material. Thus, using the optimized conditions for lithiation, followed by the
reaction with sulfur and subsequent alkylation, final compounds 31 and 35 (via crude 33,
Scheme 8) were prepared in good yields.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
9
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 10 of 27
1
2
3
4
5
6
OBn
N
Boc
N
N
24
26
Z = COOEt
Z = P(O)(OEt)2
7
8
9
EtOOC
S
Z
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
24
for : (i) TMPLi, TMEDA, THF, 2 min, -78 °C,
(ii) S8, THF, 10 min, -78 °C  0 °C
26
for : (i) LDA, THF, 2 min, -95 °C,
(ii) S8, THF, 10 min, -95 °C  0 °C
OBn
OBn
Boc
Boc
N
N
TfOCH2P(O)(Oi-Pr)2,
THF, 10 min, 0 °C
N
N
P(O)(Oi-Pr)2
N
N
S
SLi
EtOOC
EtOOC
S
S
Z
Z
25
31
Z = COOEt (68%)
Z = P(O)(OEt)2 (59%)
29
30
Z = COOEt
Z = P(O)(OEt)2
0.1M HCl (aq.), 0 °C
OBn
Boc
OBn
N
Boc
BrCH2CH2P(O)(OEt)2,
K2CO3, DMF
N
N
N
N
7 days, rt
N
EtOOC
S
SH
EtOOC
S
S
P(O)(OEt)2
Z
Z
32
33
34
35
Z = COOEt
Z = COOEt (59%)
Z = P(O)(OEt)2 (50%)
Z = P(O)(OEt)2
Scheme 8. Synthesis of target tetrasubstituted thiophenes bearing the 9-deazapurine moiety
and a linker with the phosphonate group.
CONCLUSIONS
In summary, we have developed and optimized a new strategy for the synthesis of
asymmetrically tetrasubstituted thiophenes. Direct metalation turned out to be an extremely
useful approach for regioselective halogenation of electron deficient aromatics, where
electrophilic aromatic substitution fails and/or provides a mixture of regioisomers. The
chemistry presented here afforded target polysubstituted thiophenes in high overall yields, with
low number of reaction steps, and with excellent regioselectivity control. Moreover, the
synthetic methodology may be transferable to other heterocycles in order to gain access to a
plethora of compounds with potential applications in medicinal and material chemistry.
10
ACS Paragon Plus Environment

Page 11 of 27
The Journal of Organic Chemistry
1
2
3
4
EXPERIMENTAL SECTION
5
6
7
8
General Methods. Unless otherwise stated, solvents were evaporated at 40 °C/2 kPa and
prepared compounds were dried at 30 °C at 2 kPa. Starting compounds and reagents were
purchased from commercial suppliers (Sigma-Aldrich, Fluorochem, Acros Organics,
Carbosynth, TCI) and used without further purification or were prepared according to published
procedures. Reaction flasks were heated in aluminium heating blocks. Diethyl ether,
tetrahydrofuran, dioxane, and acetonitrile were dried by activated neutral alumina
(drysphere®). Dimethylformamide was dried by activated molecular sieves (3Å). Other dry
solvents were purchased from commercial suppliers (Sigma-Aldrich, Acros Organics).
Triethylamine was dried with potassium hydroxide under argon atmosphere in dark flask sealed
with septum. Large-scale reactions were carried-out in the Syrris Atlas Potassium system with
2 l, 1 l or 0.5 l jacket reactor coupled with Julabo FP50-HL Refrigerated/Heating Circulator.
Analytical TLC was performed on silica gel pre-coated aluminium plates with fluorescent
indicator (Merck 60 F254). Flash column chromatography was carried out by Teledyne ISCO
CombiFlash Rf200 with dual absorbance detector. Various types of columns were used: a)
Teledyne ISCO columns RediSepRf HP Silica GOLD in sizes 12 g, 40 g, 80 g and 120 g; b)
Teledyne ISCO columns RediSepRf HP C18 Aq GOLD in sizes 50 g and 100 g; c) column
Chromabond Flash DL 40, DL 80, DL 120 and DL 200, filled with FLUKA silica gel 60; d)
Interchim puriFlash C18 Aq in sizes F0040 and F0080. Eluents were used: methanol (A),
cyclohexane (B), ethyl acetate modified with 10% (v/v) of methanol (C), chloroform (D), water
(E), cyclohexane modified with 0.5% of triethylamine (F). Mass spectra, UV spectra and purity
of compounds were measured on Waters UPLC-MS system consisted of Waters UPLC H-Class
Core System (column Waters Acquity UPLC BEH C18 1.7 mm, 2.1 x 100 mm), Waters
Acquity UPLC PDA detector and Mass spectrometer Waters SQD2. The universal LC method
was used (eluent H2O/ CH3CN, gradient 0 – 100%, run length 7 min) and MS method (ESI+
and/or ESI-, cone voltage = 30 V, mass detector range 100 – 1000 Da). Spectra are screened as
a PDA acquisition, absorption at 254 nm, and mass spectrum of the main peak at 254 nm.
HRMS spectra were recorded on Q-Tof micro (Waters) with CI ionization method or LTQ
Orbitrap XL spectrometer (Thermo Fisher Scientific) with ESI ionization method. NMR spectra
were recorded on Bruker Avance 400 or 500 spectrometers referenced to the residual solvent
signal or a specified additive. Assignments of NMR signals are stated in supplement data and
are based on heteronuclear correlation experiments HSQC, HMBC, COSY and NOESY in
specific cases. Melting points were measured on BÜCHI Melting Point B-545.
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
11
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 12 of 27
1
2
3
4
5
6
7
8
9
Ethyl 2-amino-5-methylthiophene-3-carboxylate (1).19 Ethyl cyanoacetate (26.6 mL,
250 mmol, 1 eq.), sulfur (8.00 g, 250 mmol, 1 eq.) and triethylamine (34.8 mL, 250 mmol, 1
eq.) were dissolved in dimethylformamide (50 mL) at room temperature. Then, a solution of
propanal (20.00 mL, 275 mmol, 1.1 eq.) in ethanol (7.50 mL) was added dropwise to the
mixture (exothermic reaction!). The reaction mixture was stirred at 60 °C and monitored by
UPLC/MS. The reaction was quenched by addition of water and extracted with ethyl acetate.
The extract was washed with water and brine, dried with magnesium sulfate and evaporated.
Purification by flash chromatography on silica gel (B to 10% of C) afforded 34 g (74%) of the
title compound as a red oil. 1H NMR (401 MHz, Chloroform-d) δ 6.59 (q, J = 1.3 Hz, 1H), 5.71
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
13
(s, 2H), 4.23 (q, J = 7.1 Hz, 2H), 2.23 (d, J = 1.3 Hz, 3H), 1.31 (t, J = 7.1 Hz, 3H). C{1H}
NMR (101 MHz, Chloroform-d) δ 165.4, 161.6, 122.6, 120.9, 106.4, 59.6, 14.9, 14.6. MS (ESI-
QMS) m/z: [M + H]+ Calcd for C8H12NO2S 186.1; Found 186.2.
Ethyl 2-bromo-5-methylthiophene-3-carboxylate (2).20 Copper (I) bromide (7.24 g, 32.4
mmol, 1.2 eq.) was suspended in dioxane (50 mL) and the mixture was cooled to 0 °C. Then,
tert-butyl nitrite (4.82 mL, 40.5 mmol, 1.5 eq.) was added and the mixture was stirred at 0 °C
for 30 minutes. Subsequently, a solution of ethyl 2-amino-5-methylthiophene-3-carboxylate (1)
(5 g, 27.0 mmol, 1 eq.) in acetonitrile (50 mL) was added and the mixture was stirred 30 minutes
at 0 °C and 2 hours at room temperature. The reaction was quenched by addition of 1M HCl
(aq.) and extracted with ethyl acetate. Extract was washed with water and brine, dried with
magnesium sulfate and evaporated. Purification by flash chromatography on silica gel (B to 5%
1
of C) afforded 3.36 g (50%) of the title compound as a yellowish oil. H NMR (401 MHz,
Chloroform-d) δ 7.01 (q, J = 1.2 Hz, 1H), 4.30 (q, J = 7.1 Hz, 2H), 2.38 (d, J = 1.2 Hz, 3H),
1.35 (t, J = 7.1 Hz, 3H). 13C{1H} NMR (101 MHz, Chloroform-d) δ 162.1, 134.0, 131.0, 127.0,
116.6, 60.9, 15.4, 14.4. MS (ESI-QMS) m/z: [M + H]+ Calcd for C8H10BrO2S 249.0; Found
249.0.
Ethyl 2-(diethoxyphosphoryl)-5-methylthiophene-3-carboxylate (3). Pd2(dba)3 (92 mg,
0.10 mmol, 0.05 eq.) and xantphos (116 mg, 0.20 mmol, 0.1 eq.) were dissolved in dioxane (50
mL), then triethylamine (419 μL, 3.00 mmol, 1.5 eq.) was added. The mixture was stirred 15
minutes at room temperature and a solution of ethyl 2-bromo-5-methylthiophene-3-carboxylate
(2) (500 mg, 2.01 mmol, 1 eq.) and diethyl phosphite (57 μL, 2.21 mmol, 1.1 eq.) in dioxane
(10 mL) was added. The mixture was stirred 2 hours at 80 °C, an then it was cooled to 0 °C,
quenched with 1M HCl (aq.), extracted with ethyl acetate, washed with brine, dried with
magnesium sulfate and evaporated. Purification by flash chromatography on silica gel (B to
20% of C) afforded 500 mg (81%) of the title compound as a yellow oil. 1H NMR (401 MHz,
12
ACS Paragon Plus Environment

Page 13 of 27
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
Chloroform-d) δ 7.27 (dd, J = 4.1, 1.0 Hz, 1H), 4.33 (q, J = 7.1 Hz, 2H), 4.27 – 4.09 (m, 4H),
2.48 (d, J = 1.0 Hz, 3H), 1.39 – 1.34 (m, 3H), 1.35 – 1.29 (m, 6H). 13C{1H} NMR (101 MHz,
Chloroform-d) δ 162.1 (d, J = 2.9 Hz), 146.8 (d, J = 7.3 Hz), 138.6 (d, J = 8.8 Hz), 131.2 (d, J
= 209.1 Hz), 129.8 (d, J = 14.6 Hz), 63.1 (d, J = 5.8 Hz), 61.3, 16.4 (d, J = 7.1 Hz), 15.2, 14.3.
31P{1H} NMR (162 MHz, Chloroform-d) δ 11.47. HRMS (CI-TOF) m/z: [M + H]+ Calcd for
C12H20O5PS 307.0769; Found 307.0770.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Ethyl 5-(bromomethyl)-2-(diethoxyphosphoryl)thiophene-3-carboxylate (4). Ethyl 2-
(diethoxyphosphoryl)-5-methylthiophene-3-carboxylate (3) (100 mg, 0.3266 mmol, 1 eq.) was
dissolved in dry acetonitrile (1 mL). Then, N-bromosuccinimide (233 mg, 1.31 mmol, 4 eq.)
was added and the mixture was stirred overnight at 80 °C. The mixture was cooled to room
temperature, quenched with water, extracted with ethyl acetate, washed with brine, dried with
magnesium sulfate and evaporated. Purification by flash chromatography on silica gel (B to
1
20% of C) afforded 100 mg (80%) of the title compound as a clear oil. H NMR (401 MHz,
Chloroform-d) δ 7.50 (d, J = 4.4 Hz, 1H), 4.56 (s, 2H), 4.27 (q, J = 7.1 Hz, 2H), 4.20 – 3.98
(m, 4H), 1.40 – 1.05 (m, 9H). 13C{1H} NMR (101 MHz, Chloroform-d) δ 161.3 (d, J = 2.9 Hz),
146.5 (d, J = 7.4 Hz), 138.0 (d, J = 8.8 Hz), 134.9 (d, J = 206.7 Hz), 131.3 (d, J = 14.5 Hz),
31
63.2 (d, J = 5.8 Hz), 61.4, 24.3, 16.2 (d, J = 6.6 Hz), 14.0. P{1H} NMR (162 MHz,
Chloroform-d) δ 10.23. HRMS (ESI-FTMS) m/z: [M + H]+ Calcd for C12H19BrO5PS 384.9869;
Found 384.9868.
Ethyl 4-bromo-2-(diethoxyphosphoryl)-5-methylthiophene-3-carboxylate (5). Ethyl 2-
(diethoxyphosphoryl)-5-methylthiophene-3-carboxylate (3) (100 mg, 0.3266 mmol, 1 eq.) was
dissolved in acetic acid (1 mL) in amber glass vial. Then, bromine (67 μL, 1.31 mmol, 4 eq.)
was added and the mixture was stirred overnight at room temperature. The mixture was cooled
to 0 °C, diluted with water, neutralized with sodium bicarbonate, extracted with ethyl acetate,
washed with saturated solution (aq.) of sodium thiosulfate, washed with brine, dried with
magnesium sulfate and evaporated. Purification by flash chromatography on silica gel (B to
1
20% of C) afforded 120 mg (96%) of the title compound a clear oil. H NMR (401 MHz,
Chloroform-d) δ 4.36 (q, J = 7.2 Hz, 2H), 4.21 – 3.96 (m, 4H), 2.41 (s, 3H), 1.36 (t, J = 7.1 Hz,
3H), 1.29 (td, J = 7.1, 0.7 Hz, 6H). 13C{1H} NMR (101 MHz, Chloroform-d) δ 163.0 (d, J = 3.1
Hz), 143.3 (d, J = 5.8 Hz), 140.2 (d, J = 10.1 Hz), 126.7 (d, J = 206.7 Hz), 110.8 (d, J = 18.4
Hz), 63.2 (d, J = 5.2 Hz), 62.2, 16.2 (d, J = 7.0 Hz), 15.4 (d, J = 1.8 Hz), 14.0. 31P{1H} NMR
(162 MHz, Chloroform-d) δ 10.12. HRMS (ESI-FTMS) m/z: [M + H]+ Calcd for C12H19BrO5PS
384.9869; Found 384.9868.
13
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 14 of 27
1
2
3
4
Tert-butyl
4-(benzyloxy)-7-(5-(diethoxyphosphoryl)-4-(ethoxycarbonyl)-2-
5
6
7
8
methylthiophen-3-yl)-5H-pyrrolo[3,2-d]pyrimidine-5-carboxylate (7). Ethyl 4-bromo-2-
(diethoxyphosphoryl)-5-methylthiophene-3-carboxylate (5) (75 mg, 0.1947 mmol, 1 eq.) was
dissolved in tetrahydrofuran (1 mL) and triethylamine (81 μL, 0.5841 mmol, 3 eq.) and 4,4,5,5-
tetramethyl-1,3,2-dioxaborolane (57 μL, 0.3894 mmol, 2 eq.) were added. The mixture was
stirred at room temperature and a solution of Pd(t-Bu3P)2 (5 mg, 0.0097 mmol, 0.05 eq.) in
tetrahydrofuran (1 mL) was added. The resulting mixture was stirred 2 hours at 40 °C, and then
filtered through syringe filter, washed with dry tetrahydrofuran and evaporated. The flask with
evaporated boronic ester was charged with tert-butyl 4-(benzyloxy)-7-bromo-5H-pyrrolo[3,2-
d]pyrimidine-5-carboxylate (9) (83 mg, 0.2044 mmol, 1.05 eq.), XPhos Pd G2 (8 mg, 0.0097
mmol, 0.05 eq.), potassium phosphate tribasic (83 mg, 0.3894 mmol, 2 eq.), and the flask was
flushed with argon. A mixture of tetrahydrofuran (4 mL) and water (1 mL) was added and the
reaction was stirred 1 hour at 40 °C. The reaction was quenched with a mixture of saturated
solution of NH4Cl (aq.) and water (1:1), extracted with ethyl acetate, washed with brine, dried
with magnesium sulfate and evaporated. Purification by flash chromatography on silica gel (F
to 20% of C) and on reverse phase (E to A) afforded 61 mg (50 %) of the title compound as a
clear oil. 1H NMR (401 MHz, Chloroform-d) δ 8.58 (s, 1H), 7.87 (s, 1H), 7.58 – 7.49 (m, 2H),
7.41 – 7.34 (m, 2H), 7.34 – 7.28 (m, 1H), 5.65 (s, 2H), 4.27 – 4.11 (m, 4H), 4.14 (q, J = 7.2 Hz,
2H), 2.42 (s, 3H), 1.53 (s, 9H), 1.35 (td, J = 7.0, 0.7 Hz, 6H), 1.15 (t, J = 7.1 Hz, 3H). 13C{1H}
NMR (101 MHz, Chloroform-d) δ 164.0 (d, J = 3.0 Hz), 156.5, 153.3, 152.3, 148.0, 146.2 (d,
J = 6.4 Hz), 141.0 (d, J = 9.7 Hz), 136.5, 131.4, 129.9 (d, J = 15.5 Hz), 128.5, 128.4 – 128.0
(m), 126.9 (d, J = 208.3 Hz), 114.5 – 113.9 (m), 85.4, 68.4, 63.1 (d, J = 5.5 Hz), 61.5, 27.9,
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
31
16.4 (d, J = 6.8 Hz), 14.7 (d, J = 2.0 Hz), 13.9. P{1H} NMR (162 MHz, Chloroform-d) δ
11.66. HRMS (ESI-FTMS) m/z: [M + H]+ Calcd for C30H37O8N3PS 630.2034; Found 630.2031.
4-(Benzyloxy)-7-bromo-5-((tert-butoxycarbonyl))-5H-pyrrolo[3,2-d]pyrimidine
(9)
The jacket reactor (2 l) was flushed with nitrogen, charged with benzyl alcohol (1 l) and the
system was set to retain temperature 20 °C. Sodium metal (22.5 g, 977 mmol, 1.5 eq.) was
added in portions and the mixture was stirred 20 hours under a small flow of nitrogen. The
mixture was heated at 80 °C for 1 hour, and then it was cooled to 20 °C. 4-Chloro-5H-
pyrrolo[3,2-d]pyrimidine (8) (100 g, 651 mmol, 1 eq.) was charged and the mixture was stirred
at 80 °C until complete conversion was achieved (ca. 4 hours). The mixture was cooled to 5 °C,
diluted with water (400 mL), pH was adjusted to pH 7 with 3M HCl (aq.) (ca. 100 - 130 mL)
and it was heated to 20 °C. The mixture was extracted with chloroform (3 x 400 mL) and the
organic phase was washed with brine (500 mL). The mixture was concentrated and benzyl
14
ACS Paragon Plus Environment

Page 15 of 27
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
alcohol was evaporated at high vacuum at ca. 90 °C. The solid was filtered through a short pad
of silica gel (600 g) with eluent (100% of D, then D with 5% of A). Solvents were evaporated
and the residue was just dissolved in a refluxed mixture of ethyl acetate/methanol (1:1) and the
solution was cooled to 30 °C. An anti-solvent pentane was slowly added (same volume as the
mixture), the mixture was cooled to -20 °C within 3 hours, and it was stirred overnight. Crystals
were collected, washed with pentane and dried. The procedure afforded 141 g (96%) of 4-
(benzyloxy)-5H-pyrrolo[3,2-d]pyrimidine as white crystals, m. p. = 157.0 – 159.0 °C. 1H NMR
(401 MHz, Chloroform-d) δ 9.33 (s, 1H), 8.58 (s, 1H), 7.49 – 7.43 (m, 2H), 7.41 (m, 1H), 7.38
– 7.32 (m, 2H), 7.35 (s, 1H), 6.67 (dd, J = 3.2, 2.1 Hz, 1H), 5.59 (s, 2H). 13C{1H} NMR (101
MHz, Chloroform-d) δ 155.5, 150.6, 150.1, 136.2, 128.8 – 128.3 (m), 115.1, 103.3, 67.9. MS
(ESI-QMS) m/z: [M + H]+ Calcd for C13H12N3O 226.1; Found 226.1. The jacket reactor (1 l)
was flushed with nitrogen and 4-(benzyloxy)-5H-pyrrolo[3,2-d]pyrimidine (40 g, 178 mmol, 1
eq.) was dissolved in tetrahydrofuran (500 mL) and N-bomosuccinimide (34.7 g, 195 mmol,
1.1 eq.) was added. The mixture was stirred at room temperature for 1 hour, during which the
product crystalized. The solvent was evaporated to dryness and the crude mixture was used in
the next step. The jacket reactor (1 l) was flushed with nitrogen and the crude mixture was
dissolved in acetonitrile (400 mL). Then, 4-(dimethylamino)pyridine (2.2 g, 18 mmol, 0.1 eq.)
was added, followed by slow addition (30 minutes) of a di-tert-butyl dicarbonate (58.2 g, 267
mmol, 1.5 eq.) solution in dry acetonitrile (100 mL). The reaction was stirred at room
temperature until gas release ceased (ca. 2 hours). Solvents were evaporated and the solid was
filtered through a short pad of neutral alumina (300 g) in a mixture of cyclohexane/ethylacetate
(3:1, v/v). Solvents were evaporated and the solid was lyophilized from dioxane to yield 58 g
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
1
(81%) of title compound 9 as a white solid, m.p. = 75.0 – 76.0 °C. H NMR (401 MHz,
Chloroform-d) δ 8.68 (s, 1H), 7.92 (s, 1H), 7.56 – 7.48 (m, 2H), 7.43 – 7.27 (m, 4H), 5.65 (s,
2H), 1.54 (s, 9H). 13C{1H} NMR (101 MHz, Chloroform-d) δ 156.5, 152.9, 152.0, 147.2, 136.3,
131.3, 128.6, 128.2, 128.1, 114.3, 96.8, 85.9, 68.7, 27.9. HRMS (ESI-FTMS) m/z: [M + H]+
Calcd for C18H19O3N3Br 404.0604; Found 404.0603.
S-(thiophen-2-yl) benzothioate (11) A flask was charged with CuI (900 mg, 4.76 mmol,
0.1 eq.) and 1,10-phenantroline (1.7 g, 9.52 mmol, 0.2 eq.) and the flask was flushed with argon.
Toluene (100 mL) was added, followed by triethylamine (9.94 mL, 71.4 mmol, 1.5 eq.). The
mixture was heated to 100 °C, then, 2-iodothiophene (10 g, 47.6 mmol, 1 eq.) and thiobenzoic
acid (6.72 mL, 57.1 mmol, 1.2 eq.) were added simultaneously at once (otherwise major
dehalogenation of 2-iodothiophene was observed). The reaction was quenched with mixture of
saturated solution of NaHCO3 (aq.) and water (1:1), extracted with ethyl acetate, washed with
15
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 16 of 27
1
2
3
4
5
6
7
8
9
brine, dried with magnesium sulfate and evaporated. Purification by flash chromatography on
silica gel (B to 2% of C) afforded 10.3 g (98%) of the title compound, which was lyophilized
from dioxane to obtain yellowish solid. 1H NMR (401 MHz, Chloroform-d) δ 8.04 (d, J = 7.7
Hz, 2H), 7.67 – 7.58 (m, 2H), 7.50 (t, J = 7.7 Hz, 2H), 7.28 (dd, J = 3.4, 1.0 Hz, 1H), 7.18 (dd,
J = 5.3, 3.6 Hz, 1H). 13C{1H} NMR (101 MHz, Chloroform-d) δ 189.8, 136.4, 136.0, 134.0,
132.2, 128.9, 128.0, 127.7, 124.2. HRMS (EI-TOF) m/z: [M]+ Calcd for C11H8OS2 220.0017;
Found 220.0019.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Ethyl 5-(benzoylthio)thiophene-2-carboxylate (12) S-(thiophen-2-yl) benzothioate (11)
(1 g, 4.54 mmol, 1 eq.) was dissolved in tetrahydrofuran (20 mL) and cooled to -50 °C. A
solution of 2,2,6,6-tetramethylpiperidinylmagnesium chloride lithium chloride complex (9.99
mL, 9.99 mmol, 2.2 eq., 1M) in tetrahydrofuran was added dropwise and the mixture was stirred
30 minutes at -50 °C. Then, ethyl cyanoformate (989 μL, 9.99 mmol, 2.2 eq.) was added and
the resulting mixture was stirred 30 minutes at -25 °C. The reaction was quenched with a
mixture of saturated solution of NH4Cl (aq.) and water (1:1), extracted with ethyl acetate,
washed with brine, dried with magnesium sulfate and evaporated. Purification by flash
chromatography on silica gel (B to 10% of C) afforded 1.31 g (99%) of the title compound as
a red oil. 1H NMR (401 MHz, Chloroform-d) δ 7.97 (dd, J = 8.4, 1.3 Hz, 2H, 9, 13), 7.78 (d, J
= 3.9 Hz, 1H, 3), 7.63 – 7.54 (m, 1H, 11), 7.51 – 7.42 (m, 2H, 10, 12), 7.19 (d, J = 3.9 Hz, 1H,
13
4), 4.35 (q, J = 7.2 Hz, 2H, 18), 1.36 (t, J = 7.2 Hz, 3H, 19). C{1H} NMR (101 MHz,
Chloroform-d) δ 188.0, 161.3, 138.7, 135.5, 135.3, 134.2, 133.1, 132.2, 128.9, 127.6, 61.4,
14.3. HRMS (EI-TOF) m/z: [M]+ Calcd for C14H12O3S2 292.0230; Found 292.0228.
Ethyl 4-bromothiophene-3-carboxylate (15).30 3,4-Dibromothiophene (14) (10 g, 41.3
mmol, 1 eq.) was dissolved in tetrahydrofuran (100 mL) and cooled to 0 °C. A solution of
isopropylmagnesium bromide (19.1 mL, 51.7 mmol, 1.25 eq., 2.7M) was added to the mixture
keeping the temperature below 5 °C and the mixture was stirred 5 hours at 0 °C. Then, ethyl
chloroformate (7.9 mL, 82.7 mmol, 2 eq.) was added keeping the temperature below 5 °C. The
resulting mixture was stirred 15 minutes at 0 °C and overnight at room temperature. The
reaction was cooled to 0 °C, quenched with 1M HCl (aq.), extracted with ethyl acetate, washed
with brine, dried with magnesium sulfate and evaporated. Purification by flash chromatography
1
on silica gel (B to 10% of C) afforded 8.0 g (82%) of the title compound as a yellow oil. H
NMR (401 MHz, Chloroform-d) δ 8.07 (d, J = 3.7 Hz, 1H), 7.27 (d, J = 3.6 Hz, 1H), 4.31 (q, J
= 7.1 Hz, 2H), 1.34 (t, J = 7.1 Hz, 3H).13C{1H} NMR (101 MHz, Chloroform-d) δ 161.3, 134.2,
131.3, 125.3, 110.8, 61.0, 14.3. MS (ESI-QMS) m/z: [M + H]+ Calcd for C7H8BrO2S 234.9;
Found 235.0.
16
ACS Paragon Plus Environment

Page 17 of 27
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
Diethyl 4-bromothiophene-2,3-dicarboxylate (16). Ethyl 4-bromothiophene-3-
carboxylate (15) (5 g, 21.3 mmol, 1 eq.) was dissolved in tetrahydrofuran (50 mL) and cooled
to -20 °C. A solution of 2,2,6,6-tetramethylpiperidinylmagnesium chloride lithium chloride
complex (25.5 mL, 25.5 mmol, 1.2 eq., 1M) in tetrahydrofuran was added dropwise and the
mixture was stirred 30 minutes at -20 °C. Then, ethyl cyanoformate (3.6 mL, 31.9 mmol, 1.5
eq.) was added and the resulting mixture was stirred 30 minutes at 0 °C. The reaction was
quenched with mixture of saturated solution of NH4Cl (aq.) and water (1:1), extracted with
ethyl acetate, washed with brine, dried with magnesium sulfate and evaporated. Purification by
flash chromatography on silica gel (B to 10% of C) afforded 5.79 g (72%) of the title compound
as a clear oil. 1H NMR (400 MHz, DMSO-d6) δ 8.16 (s, 1H), 4.41 – 4.23 (m, 4H), 1.30 (t, J =
7.1 Hz, 3H), 1.26 (t, J = 7.1 Hz, 3H). 13C{1H} NMR (101 MHz, DMSO-d6) δ 163.2, 159.3,
139.2, 131.6, 131.3, 108.2, 62.1, 62.0, 13.9. HRMS (CI-TOF) m/z: [M + H]+ Calcd for
C10H12BrO4S 306.9640; Found 306.9641.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Diethyl
4-bromo-5-(((diisopropoxyphosphoryl)methyl)thio)thiophene-2,3-
dicarboxylate (17). Diethyl 4-bromothiophene-2,3-dicarboxylate (16) (1 g, 3.26 mmol, 1 eq.)
was dissolved in tetrahydrofuran (20 mL) and cooled to -20 °C. A solution of 2,2,6,6-
tetramethylpiperidinylmagnesium chloride lithium chloride complex (3.9 mL, 3.91 mmol, 1.2
eq., 1M) in tetrahydrofuran was added dropwise and the mixture was stirred 30 minutes at
-20 °C. Then, a solution of sulfur (156 mg, 4.89 mmol, 1.5 eq.) in tetrahydrofuran (10 mL) was
added and the resulting mixture was stirred at 0 °C. After 30 minutes, diisopropyl
triflyloxymethylphosphonate (1.46 g, 4.89 mmol, 1.5 eq.) was added and the mixture was
stirred additional 30 minutes at 0 °C. The reaction was quenched with mixture of saturated
solution of NH4Cl (aq.) and water (1:1), extracted with ethyl acetate, washed with brine, dried
with magnesium sulfate and evaporated. Purification by flash chromatography on silica gel (B
to 10% of C) afforded 1.65 g (98%) of the title compound as a yellow oil. 1H NMR (401 MHz,
Chloroform-d) δ 4.75 (dhept, J = 7.7, 6.2 Hz, 2H), 4.42 (q, J = 7.2 Hz, 2H), 4.32 (q, J = 7.1 Hz,
2H), 3.17 (d, J = 14.0 Hz, 2H), 1.39 (t, J = 7.1 Hz, 3H), 1.36 – 1.30 (m, 15H). 13C{1H} NMR
(101 MHz, Chloroform-d) δ 163.6, 159.5, 140.4, 138.9 (d, J = 8.0 Hz), 131.9, 114.9, 72.0 (d, J
= 6.9 Hz), 62.6, 62.3, 31.2 (d, J = 147.9 Hz), 26.5 – 22.4 (m), 14.2, 14.1. HRMS (ESI-FTMS)
m/z: [M + H]+ Calcd for C17H27O7BrPS2 517.0114; Found 517.0107.
Ethyl 4-bromo-2-(diethoxyphosphoryl)thiophene-3-carboxylate (18). Pd2(dba)3 (729
mg, 0.80 mmol, 0.05 eq.) and xantphos (922 mg, 1.59 mmol, 0.1 eq.) were dissolved in dioxane
(100 mL) and triethylamine (3.32 mL, 23.9 mmol, 1.5 eq.) was added. The mixture was stirred
15 minutes at room temperature and a solution of ethyl 2,4-dibromothiophene-3-carboxylate
17
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 18 of 27
1
2
3
4
5
6
7
8
9
(20) (5 g, 15.9 mmol, 1 eq.) and diethyl phosphite (2.25 mL, 17.5 mmol, 1.1 eq.) in dioxane (20
mL) was added. The mixture was stirred 2 hours at 80 °C, then it was cooled to 0 °C, quenched
with 1M HCl (aq.), extracted with ethyl acetate, washed with brine, dried with magnesium
sulfate and evaporated. Purification by flash chromatography on silica gel (B to 20% of C)
afforded 5.04 g (85%) of the title compound as a yellow oil. 1H NMR (401 MHz, Chloroform-
d) δ 7.57 (d, J = 6.0 Hz, 1H), 4.41 (q, J = 7.2 Hz, 2H), 4.26 – 4.05 (m, 5H), 1.40 (t, J = 7.1 Hz,
3H), 1.37 – 1.29 (m, 6H). 13C{1H} NMR (101 MHz, Chloroform-d) δ 162.7 (d, J = 2.9 Hz),
139.8 (d, J = 10.3 Hz), 131.8 (d, J = 204.0 Hz), 131.1 (d, J = 5.8 Hz), 111.6 (d, J = 19.7 Hz),
63.6 – 63.2 (m), 62.4, 16.8 – 16.0 (m), 14.1. 31P{1H} NMR (162 MHz, Chloroform-d) δ 9.46.
HRMS (CI-TOF) m/z: [M + H]+ Calcd for C11H17BrO5PS 370.9718; Found 370.9717.
Ethyl 4-bromo-2-iodothiophene-3-carboxylate (19). Ethyl 4-bromothiophene-3-
carboxylate (15) (5 g, 21.3 mmol, 1 eq.) was dissolved in tetrahydrofuran (50 mL) and cooled
to -20 °C. A solution of 2,2,6,6-tetramethylpiperidinylmagnesium chloride lithium chloride
complex (25.5 mL, 25.52 mmol, 1.2 eq., 1M) in tetrahydrofuran was added dropwise and the
mixture was stirred 30 minutes at -20 °C. Then, a solution of iodine (8.1 g, 31.91 mmol, 1.5
eq.) in tetrahydrofuran (30 mL) was added and the resulting mixture was stirred 30 minutes at
0 °C. The reaction was quenched with mixture of saturated solution of NH4Cl (aq.) and water
(1:1), extracted with ethyl acetate, washed with brine, dried with magnesium sulfate and
evaporated. Purification by flash chromatography on silica gel (B to 10% of C) afforded 6.8 g
(88%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 7.41 (s, 1H),
4.40 (q, J = 7.2 Hz, 2H), 1.41 (t, J = 7.2 Hz, 3H). 13C{1H} NMR (101 MHz, Chloroform-d) δ
162.1, 136.6, 130.5, 108.9, 79.6, 61.9, 14.3. HRMS (CI-TOF) m/z: [M + H]+ Calcd for
C7H7BrIO2S 360.8395; Found 360.8395.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Ethyl 2,4-dibromothiophene-3-carboxylate (20). Ethyl 4-bromothiophene-3-
carboxylate (15) (5 g, 21.3 mmol, 1 eq.) was dissolved in tetrahydrofuran (50 mL) and cooled
to -20 °C. A solution of 2,2,6,6-tetramethylpiperidinylmagnesium chloride lithium chloride
complex (25.5 mL, 25.52 mmol, 1.2 eq., 1M) in tetrahydrofuran was added dropwise and the
mixture was stirred 30 minutes at -20 °C. Then, a solution of 1,2-dibromotetrachloroethane
(10.4 g, 31.9 mmol, 1.5 eq.) in tetrahydrofuran (30 mL) was added and the resulting mixture
was stirred 30 minutes at 0 °C. The reaction was quenched with mixture of saturated solution
of NH4Cl (aq.) and water (1:1), extracted with ethyl acetate, washed with brine, dried with
magnesium sulfate and evaporated. Purification by flash chromatography on silica gel (B to
10% of C) afforded 5.4 g (81%) of the title compound as a yellowish oil. 1H NMR (401 MHz,
Chloroform-d) δ 7.27 (s, 1H), 4.41 (q, J = 7.1 Hz, 2H), 1.41 (t, J = 7.1 Hz, 3H). 13C{1H} NMR
18
ACS Paragon Plus Environment

Page 19 of 27
The Journal of Organic Chemistry
1
2
3
4
5
6
(101 MHz, Chloroform-d) δ 161.8, 132.5, 125.2, 117.3, 109.6, 62.0, 14.3. HRMS (CI-TOF)
m/z: [M + H]+ Calcd for C7H7Br2O2S 312.8533; Found 312.8532.
Diethyl
4-(4-(benzyloxy)-5-(tert-butoxycarbonyl)-5H-pyrrolo[3,2-d]pyrimidin-7-
7
8
9
yl)thiophene-2,3-dicarboxylate (24). Diethyl 4-bromothiophene-2,3-dicarboxylate (16) (3 g,
9.77 mmol, 1 eq.) was dissolved in tetrahydrofuran (30 mL) and triethylamine (4.1 mL, 29.3
mmol, 3 eq.) and 4,4,5,5-tetramethyl-1,3,2-dioxaborolane (2.8 mL, 19.5 mmol, 2 eq.) were
added. The mixture was stirred at room temperature and a solution of Pd(t-Bu3P)2 (250 mg,
0.49 mmol, 0.05 eq.) in tetrahydrofuran (5 mL) was added. The resulting mixture was stirred 2
hours at 40 °C, and then filtered through celite, washed with dry tetrahydrofuran and
evaporated. The flask with evaporated boronic ester was charged with tert-butyl 4-(benzyloxy)-
7-bromo-5H-pyrrolo[3,2-d]pyrimidine-5-carboxylate (9) (4.1 g, 10.3 mmol, 1.05 eq.), XPhos
Pd G2 (231 mg, 0.29 mmol, 0.03 eq.), potassium phosphate tribasic (4.1 g, 19.54 mmol, 2 eq.),
and the flask was flushed with argon. A mixture of tetrahydrofuran (60 mL) and water (15 mL)
was added and the reaction was stirred 1 hour at 40 °C. The reaction was quenched with a
mixture of saturated solution of NH4Cl (aq.) and water (1:1), extracted with ethyl acetate,
washed with brine, dried with magnesium sulfate and evaporated. Purification by flash
chromatography on silica gel (F to 20% of C) and on reverse phase (E to A) afforded 3.37 g
(82%) of the title compound as a yellowish solid, which was lyophilized from dioxane. 1H NMR
(500 MHz, DMSO-d6) δ 8.67 (s, 1H), 8.61 (s, 1H), 7.98 (s, 1H), 7.53 (d, J = 7.3 Hz, 2H), 7.38
(t, J = 7.4 Hz, 2H), 7.32 (t, J = 7.3 Hz, 1H), 5.63 (s, 2H), 4.44 – 4.18 (m, 4H), 1.48 (s, 9H), 1.31
– 1.25 (m, 6H).13C{1H} NMR (126 MHz, DMSO-d6) δ 165.4, 160.3, 155.9, 152.3, 151.8, 147.2,
137.4, 136.3, 131.6, 131.0, 129.6, 128.6, 128.4, 128.0, 127.8, 113.6, 112.7, 85.6, 67.9, 61.9,
61.7, 27.2, 14.0, 13.8. HRMS (ESI-FTMS) m/z: [M + H]+ Calcd for C28H30N3O7S 552.1799;
Found 552.1792.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Diethyl 4-(4-(benzyloxy)-5-(tert-butoxycarbonyl)-5H-pyrrolo[3,2-d]pyrimidin-7-yl)-5-
(((diisopropoxyphosphoryl)methyl)thio)thiophene-2,3-dicarboxylate (25). A flask was charged
with tetrahydrofuran (2 mL), 2,2,6,6-tetramethylpiperidine (370 μL, 2.18 mmol, 1.2 eq.),
N,N,N′,N′-tetramethylethylenediamine (326 μL, 2.18 mmol, 1.2 eq.) and the mixture was stirred
and cooled to -78 °C. Then, n-butyllithium (870 μL, 2.18 mmol, 1.2 eq., 2.5M) was added
dropwise and the mixture was stirred for 5 minutes at -78 °C. This solution was added during
30 seconds to a solution of diethyl 4-(4-(benzyloxy)-5-(tert-butoxycarbonyl)-5H-pyrrolo[3,2-
d]pyrimidin-7-yl)thiophene-2,3-dicarboxylate (24) (1 g, 1.81 mmol, 1 eq.) in tetrahydrofuran
(20 mL) at -78 °C. After 2 minutes of stirring, a room temperature solution of sulfur (116 mg,
3.62 mmol, 2 eq.) in tetrahydrofuran (15 mL) was added at once. The resulting mixture was
19
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 20 of 27
1
2
3
4
5
6
7
8
9
stirred 2 minutes at -78 °C, and then 10 minutes at 0 °C. Then, diisopropyl
triflyloxymethylphosphonate (892 mg, 2.72 mmol, 1.5 eq.) was added and the mixture was
stirred for 30 minutes at 0 °C. The reaction was quenched with a mixture of saturated solution
of NH4Cl (aq.) and water (1:1), extracted with ethyl acetate, washed with brine, dried with
magnesium sulfate and evaporated. Purification by flash chromatography on silica gel (F to
20% of C) afforded 939 mg (68%) of the title compound as a yellow oil. 1H NMR (401 MHz,
Chloroform-d) δ 8.54 (s, 1H), 7.96 (s, 1H), 7.53 – 7.43 (m, 2H), 7.36 – 7.29 (m, 2H), 7.29 –
7.23 (m, 1H), 5.61 (s, 2H), 4.60 (dhept, J = 7.8, 6.2 Hz, 2H), 4.28 (q, J = 7.1 Hz, 2H), 4.14 (q,
J = 7.2 Hz, 2H), 3.13 (d, J = 14.4 Hz, 2H), 1.49 (s, 9H), 1.29 (t, J = 7.2 Hz, 4H), 1.22 (d, J =
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
13
6.2 Hz, 6H), 1.17 (d, J = 6.0 Hz, 6H), 1.15 (t, J = 7.1 Hz, 3H). C{1H} NMR (101 MHz,
Chloroform-d) δ 164.4, 159.9, 156.2, 152.8, 152.0, 147.5, 141.0 (d, J = 8.9 Hz), 139.6, 136.2,
133.5, 132.3, 131.8, 128.3, 127.9, 113.9, 112.5, 85.3, 71.5 (d, J = 6.6 Hz), 68.1, 61.6, 32.9 (d,
J = 146.3 Hz), 27.6, 23.8 (d, J = 4.2 Hz), 23.6 (d, J = 4.6 Hz), 14.0, 13.7. 31P{1H} NMR (162
MHz, Chloroform-d) δ 21.48. HRMS (ESI-FTMS) m/z: [M + H]+ Calcd for C35H45N3O10PS2
762.2279; Found 762.2276.
Tert-butyl 4-(benzyloxy)-7-(5-(diethoxyphosphoryl)-4-(ethoxycarbonyl)thiophen-3-yl)-
5H-pyrrolo[3,2-d]pyrimidine-5-carboxylate
(26).
Ethyl
4-bromo-2-
(diethoxyphosphoryl)thiophene-3-carboxylate (18) (1 g, 2.69 mmol, 1 eq.) was dissolved in
tetrahydrofuran (10 mL) and triethylamine (1.1 mL, 8.08 mmol, 3 eq.) and 4,4,5,5-tetramethyl-
1,3,2-dioxaborolane (780 μL, 5.39 mmol, 2 eq.) were added. The mixture was stirred at room
temperature and a solution of Pd(t-Bu3P)2 (69 mg, 0.13 mmol, 0.05 eq.) in tetrahydrofuran (2
mL) was added. Resulting mixture was stirred 2 hours at 40 °C, and then filtered through celite,
washed with dry tetrahydrofuran and evaporated. The flask with evaporated boronic ester was
charged with tert-butyl 4-(benzyloxy)-7-bromo-5H-pyrrolo[3,2-d]pyrimidine-5-carboxylate
(9) (1.14 g, 2.82 mmol, 1.05 eq.), XPhos Pd G2 (64 mg, 0.08 mmol, 0.03 eq.), potassium
phosphate tribasic (1.1 g, 5.34 mmol, 2 eq.), and the flask was flushed with argon. A mixture
of tetrahydrofuran (20 mL) and water (5 mL) was added and the reaction was stirred 1 hour at
40 °C. The reaction was quenched with mixture of saturated solution of NH4Cl (aq.) and water
(1:1), extracted with ethyl acetate, washed with brine, dried with magnesium sulfate and
evaporated. Purification by flash chromatography on silica gel (F to 20% of C) afforded 1.26 g
(76%) of the title compound as a clear oil. 1H NMR (401 MHz, Chloroform-d) δ 8.64 (s, 1H),
8.33 (d, J = 6.1 Hz, 1H), 8.04 (s, 1H), 7.56 – 7.49 (m, 2H), 7.41 – 7.29 (m, 3H), 5.66 (s, 2H),
4.34 (q, J = 7.2 Hz, 2H), 4.29 – 4.08 (m, 4H), 1.54 (s, 9H), 1.41 – 1.27 (m, 9H). 13C{1H} NMR
(101 MHz, Chloroform-d) δ 165.10 (d, J = 2.9 Hz), 156.6, 152.9, 152.4, 147.9, 139.6 (d, J =
20
ACS Paragon Plus Environment

Page 21 of 27
The Journal of Organic Chemistry
1
2
3
4
5
6
7
8
9
11.1 Hz), 136.5, 132.5 (d, J = 16.9 Hz), 132.0 (d, J = 6.5 Hz), 131.4, 130.2 (d, J = 204.7 Hz),
129.8, 128.6, 128.2, 128.1, 114.5, 114.1, 85.1, 68.4, 63.4 – 63.0 (m), 62.2, 27.9, 16.7 – 15.9
(m), 14.1. 31P{1H} NMR (162 MHz, Chloroform-d) δ 11.47. HRMS (ESI-FTMS) m/z: [M +
H]+ Calcd for C29H35N3O8PS 616.1877; Found 616.1876.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Diethyl 4-(4-(benzyloxy)-5-(tert-butoxycarbonyl)-5H-pyrrolo[3,2-d]pyrimidin-7-yl)-5-
iodothiophene-2,3-dicarboxylate (28). A flask was charged with tetrahydrofuran (2 mL),
2,2,6,6-tetramethylpiperidine
(370
μL,
2.18
mmol,
1.2
eq.),
N,N,N′,N′-
tetramethylethylenediamine (326 μL, 2.18 mmol, 1.2 eq.) and the mixture was stirred and
cooled to -78 °C. Then, n-butyllithium (870 μL, 2.18 mmol, 1.2 eq., 2.5M) was added dropwise
and the mixture was stirred for 5 minutes at -78 °C. This solution was added during 30 seconds
to a solution of diethyl 4-(4-(benzyloxy)-5-(tert-butoxycarbonyl)-5H-pyrrolo[3,2-d]pyrimidin-
7-yl)thiophene-2,3-dicarboxylate (24) (1 g, 1.81 mmol, 1 eq.) in tetrahydrofuran (20 mL) at -
78 °C. After 2 minutes of stirring, a room temperature solution of iodine (919 mg, 3.62 mmol,
2 eq.) in tetrahydrofuran (15 mL) was added at once. The resulting mixture was stirred 2
minutes at -78 °C and then 10 minutes at 0 °C. The reaction was quenched with a mixture of
saturated solution of NH4Cl (aq.) and water (1:1), extracted with ethyl acetate, washed with a
saturated aqueous solution of Na2S2O5, washed with brine, dried with magnesium sulfate and
evaporated. Purification by flash chromatography on silica gel (F to 20% of C) afforded 882
mg (72%) of the title compound as a yellow oil. 1H NMR (500 MHz, DMSO-d6) δ 8.58 (s, 1H),
8.15 (s, 1H), 7.59 – 7.53 (m, 2H), 7.43 – 7.36 (m, 2H), 7.38 – 7.31 (m, 1H), 5.64 (s, 2H), 4.29
(q, J = 7.1 Hz, 2H), 4.07 (q, J = 7.1 Hz, 2H), 1.49 (s, 9H), 1.27 (t, J = 7.1 Hz, 3H), 1.04 (t, J =
7.1 Hz, 3H). 13C{1H} NMR (126 MHz, DMSO-d6) δ 162.9, 159.4, 155.9, 152.0, 151.9, 147.3,
139.1, 136.9, 136.3, 135.5, 132.2, 128.4, 128.0, 128.0, 113.7, 113.1, 91.6, 85.8, 68.0, 61.9, 61.5,
27.1, 13.9, 13.5. HRMS (ESI-FTMS) m/z: [M + H]+ Calcd for C28H29IN3O7S 678.0771; Found
678.0770 [M + H]+.
Tert-butyl
4-(benzyloxy)-7-(5-(diethoxyphosphoryl)-2-
(((diisopropoxyphosphoryl)methyl)thio)-4-(ethoxycarbonyl)thiophen-3-yl)-5H-pyrrolo[3,2-
d]pyrimidine-5-carboxylate (31). Tert-butyl 4-(benzyloxy)-7-(5-(diethoxyphosphoryl)-4-
(ethoxycarbonyl)thiophen-3-yl)-5H-pyrrolo[3,2-d]pyrimidine-5-carboxylate (26) (1 g, 1.62
mmol, 1 eq.) was dissolved in tetrahydrofuran (20 mL) and the solution was cooled to -95 °C
in bath containing methanol and liquid nitrogen. Then, a solution of lithium diisopropylamide
(2.43 mL, 2.43 mmol, 1.5 eq., 1M) was added dropwise and the mixture was stirred 5 minutes.
Then, a room temperature solution of sulfur (104 mg, 3.24 mmol, 2 eq.) in tetrahydrofuran (10
mL) was added at once. The resulting mixture was stirred 2 minutes at -95 °C, and then 10
21
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 22 of 27
1
2
3
4
5
6
7
8
9
minutes at 0 °C. Then, diisopropyl triflyloxymethylphosphonate (797 mg, 2.72 mmol, 1.5 eq.)
was added and the mixture was stirred additional 30 minutes at 0 °C. The reaction was quenched
with mixture of saturated solution of NH4Cl (aq.) and water (1:1), extracted with ethyl acetate,
washed with brine, dried with magnesium sulfate and evaporated. Purification by flash
chromatography on silica gel (F to 20% of C) afforded 843 g (59%) of the title compound as a
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
1
yellow oil. H NMR (401 MHz, Chloroform-d) δ 8.45 (s, 1H), 7.96 (s, 1H), 7.47 – 7.39 (m,
2H), 7.30 – 7.22 (m, 2H), 7.24 – 7.13 (m, 1H), 5.55 (s, 2H), 4.54 (hept, J = 7.7, 6.2 Hz, 2H),
4.19 – 4.07 (m, 4H), 4.02 (q, J = 7.1 Hz, 2H), 3.04 (d, J = 14.3 Hz, 2H), 1.43 (s, 9H), 1.26 (td,
J = 7.1, 0.6 Hz, 6H), 1.16 (d, J = 6.2 Hz, 6H), 1.11 (d, J = 6.2 Hz, 6H), 1.03 (t, J = 7.1 Hz, 3H).
13C{1H} NMR (101 MHz, Chloroform-d) δ 162.9 (d, J = 3.5 Hz), 156.1, 152.9, 151.8, 147.5,
141.3 (dd, J = 9.0, 6.2 Hz), 140.4 (d, J = 9.5 Hz), 136.1, 135.1 (d, J = 14.7 Hz), 132.2 (d, J =
205.1 Hz), 131.8, 128.2, 127.8, 113.7, 113.0, 85.1, 71.4 (d, J = 6.6 Hz), 68.0, 62.9 (d, J = 5.3
Hz), 61.3, 32.6 (d, J = 146.0 Hz), 27.5, 23.6 (dd, J = 14.9, 4.6 Hz), 16.0 (d, J = 6.7 Hz), 13.4.
31P{1H} NMR (162 MHz, Chloroform-d) δ 21.35, 9.92. HRMS (ESI-FTMS) m/z: [M + H]+
Calcd for C36H50N3O11P2S2 826.2357; Found 826.2351.
Diethyl 4-(4-(benzyloxy)-5-(tert-butoxycarbonyl)-5H-pyrrolo[3,2-d]pyrimidin-7-yl)-5-
((2-(diethoxyphosphoryl)ethyl)thio)thiophene-2,3-dicarboxylate (34). Followed the previous
procedure, after the addition of the sulfur solution and 10 minutes of stirring, the reaction was
quench with 0.1M HCl (aq.) and extracted with ethyl acetate, washed with brine, dried with
magnesium sulfate and evaporated. The crude solid was then dissolved in N,N-
dimethylformamide (30 mL) and potassium carbonate (749 mg, 5.43 mmol, 3 eq.) was added,
followed by addition of diethyl 2-bromoethylphosphonate (1.3 mL, 7.24 mmol, 4 eq.). The
mixture was stirred 7 days at room temperature. The reaction was quenched with mixture of
saturated solution of NH4Cl (aq.) and water (1:1), extracted with ethyl acetate, washed with
brine, dried with magnesium sulfate and evaporated. Purification by flash chromatography on
1
silica gel (F to 20% of C) afforded 800 mg (59%) of the title compound as a yellow oil. H
NMR (401 MHz, Chloroform-d) δ 8.55 (d, J = 0.8 Hz, 1H), 7.92 (d, J = 0.6 Hz, 1H), 7.55 –
7.44 (m, 2H), 7.38 – 7.32 (m, 2H), 7.32 – 7.27 (m, 1H), 5.61 (s, 2H), 4.30 (qt, J = 7.1, 1.0 Hz,
2H), 4.21 – 4.11 (m, 2H), 4.07 – 3.91 (m, 4H), 3.10 – 2.98 (m, 2H), 2.07 – 1.88 (m, 2H), 1.59
– 1.43 (m, 9H), 1.31 (t, J = 7.2 Hz, 3H), 1.24 (t, J = 7.1 Hz, 6H), 1.17 (t, J = 7.1 Hz, 3H).
13C{1H} NMR (101 MHz, Chloroform-d) δ 164.6, 160.1, 156.3, 152.9, 152.1, 147.7, 140.7,
139.9, 136.3, 133.2, 132.1, 131.8, 128.4, 128.0, 114.0, 112.7, 85.5, 68.3, 61.9 (d, J = 6.6 Hz),
61.8, 61.8, 30.9 (d, J = 3.2 Hz), 27.7, 26.4 (d, J = 137.0 Hz), 16.4 (d, J = 6.2 Hz), 14.2, 13.8.
22
ACS Paragon Plus Environment

Page 23 of 27
The Journal of Organic Chemistry
1
2
3
4
31P{1H} NMR (162 MHz, Chloroform-d) δ 29.94. HRMS (ESI-FTMS) m/z: [M + H]+ Calcd
5
6
7
for C34H43N3O10PS2 748.2122; Found 748.2120.
Tert-butyl
4-(benzyloxy)-7-(5-(diethoxyphosphoryl)-2-((2-
8
9
(diethoxyphosphoryl)ethyl)thio)-4-(ethoxycarbonyl)thiophen-3-yl)-5H-pyrrolo[3,2-
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
d]pyrimidine-5-carboxylate (35). Followed the previous procedure, after the addition of the
sulfur solution and 10 minutes of stirring, the reaction was quench with 0.1M HCl (aq.) and
extracted with ethyl acetate, washed with brine, dried with magnesium sulfate and evaporated.
The crude solid was then dissolved in N,N-dimethylformamide (30 mL) and potassium
carbonate (671 mg, 4.86 mmol, 3 eq.) was added, followed by addition of diethyl 2-
bromoethylphosphonate (1.2 mL, 6.48 mmol, 4 eq.). The mixture was stirred 3 days at room
temperature. The reaction was quenched with mixture of saturated solution of NH4Cl (aq.) and
water (1:1), extracted with ethyl acetate, washed with brine, dried with magnesium sulfate and
evaporated. Purification by flash chromatography on silica gel (F to 20% of C) afforded 658
mg (50%) of the title compound as a yellow oil. 1H NMR (401 MHz, Chloroform-d) δ 8.54 (s,
1H), 7.96 (s, 1H), 7.58 – 7.46 (m, 2H), 7.40 – 7.33 (m, 2H), 7.33 – 7.27 (m, 1H), 5.63 (s, 2H),
4.29 – 4.15 (m, 4H), 4.12 (q, J = 7.1 Hz, 2H), 4.07 – 3.94 (m, 4H), 3.11 – 2.93 (m, 2H), 2.03 –
1.89 (m, 2H), 1.52 (s, 9H), 1.35 (td, J = 7.1, 0.7 Hz, 6H), 1.25 (t, J = 7.1 Hz, 6H), 1.13 (t, J =
7.2 Hz, 3H). 13C{1H} NMR (101 MHz, Chloroform-d) δ 163.4 (d, J = 3.0 Hz), 156.5, 153.2,
152.1, 147.9, 141.2 (d, J = 6.1 Hz), 140.8 (d, J = 9.4 Hz), 136.5, 135.2 (d, J = 14.7 Hz), 132.4
(d, J = 205.4 Hz), 132.1, 128.5, 128.1, 114.1, 113.4, 85.5, 68.4, 63.4 (d, J = 5.3 Hz), 62.0 (d, J
= 6.6 Hz), 61.7, 30.9 (d, J = 3.2 Hz), 27.8, 26.5 (d, J = 137.1 Hz), 16.5 (d, J = 5.9 Hz), 16.4 (d,
J = 7.3 Hz), 13.8. 31P{1H} NMR (162 MHz, Chloroform-d) δ 29.93, 10.14. HRMS (ESI-FTMS)
m/z: [M + H]+ Calcd for C35H48N3O11P2S2 812.2200; Found 812.2198.
ASSOCIATED CONTENT
Supporting Information. Copies of the NMR (1H, 13C{1H}, 31P{1H}) and UPLC/MS spectra
with full signal assignment.
AUTHOR INFORMATION
Corresponding author
E-mail: janeba@uochb.cas.cz
Orcid no.
Zlatko Janeba: 0000-0003-4654-679X
Martin Dračínský: 0000-0002-4495-0070
23
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 24 of 27
1
2
3
4
Notes
5
6
The authors declare no conflict of interest.
7
8
9
ACKNOWLEDGMENTS
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
This work was supported by the Institute of Organic Chemistry and Biochemistry (RVO
61388963) and by the Technology Agency of the Czech Republic (Program NCK1, no.
TN01000013).
REFERENCES
(1)
Baszczyňski, O.; Kaiser, M. M.; Česnek, M.; Břehová, P.; Jansa, P.; Procházková, E.;
Dračínský, M.; Snoeck, R.; Andrei, G.; Janeba, Z. Xanthine-Based Acyclic Nucleoside
Phosphonates with Potent Antiviral Activity against Varicella-Zoster Virus and Human
Cytomegalovirus. Antivir. Chem. Chemother. 2018, 26, 1–13.
(2)
Klejch, T.; Pohl, R.; Janeba, Z.; Sun, M.; Keough, D. T.; Guddat, L. W.; Hocková, D.
Acyclic Nucleoside Phosphonates with Unnatural Nucleobases, Favipiravir and
Allopurinol, Designed as Potential Inhibitors of the Human and Plasmodium
Falciparum 6-Oxopurine Phosphoribosyltransferases. Tetrahedron 2018, 74, 5886–
5897.
(3)
(4)
(5)
(6)
Doleželová, E.; Terán, D.; Gahura, O.; Kotrbová, Z.; Procházková, M.; Keough, D.;
Špaček, P.; Hocková, D.; Guddat, L.; Zíková, A. Evaluation of the Trypanosoma
Brucei 6-Oxopurine Salvage Pathway as a Potential Target for Drug Discovery. PLoS
Negl. Trop. Dis. 2018, 12, e0006301.
Keough, D. T.; Rejman, D.; Pohl, R.; Zborníková, E.; Hocková, D.; Croll, T.; Edstein,
M. D.; Birrell, G. W.; Chavchich, M.; Naesens, L. M. J.; et al. Design of Plasmodium
Vivax Hypoxanthine-Guanine Phosphoribosyltransferase Inhibitors as Potential
Antimalarial Therapeutics. ACS Chem. Biol. 2018, 13, 82–90.
Lukáč, M.; Hocková, D.; Keough, D. T.; Guddat, L. W.; Janeba, Z. Novel Nucleotide
Analogues Bearing (1H-1,2,3-Triazol-4-Yl)Phosphonic Acid Moiety as Inhibitors of
Plasmodium and Human 6-Oxopurine Phosphoribosyltransferases. Tetrahedron 2017,
73, 692–702.
Špaček, P.; Keough, D. T.; Chavchich, M.; Dračínský, M.; Janeba, Z.; Naesens, L.;
Edstein, M. D.; Guddat, L. W.; Hocková, D. Synthesis and Evaluation of Symmetric
24
ACS Paragon Plus Environment

Page 25 of 27
The Journal of Organic Chemistry
1
2
3
4
5
6
Acyclic Nucleoside Bisphosphonates as Inhibitors of the Plasmodium Falciparum,
Plasmodium Vivax and Human 6-Oxopurine Phosphoribosyltransferases and the
Antimalarial Activity of Their Prodrugs. Bioorg. Med. Chem. 2017, 25, 4008–4030.
7
8
9
(7)
(8)
(9)
Kaiser, M. M.; Baszczyňski, O.; Hocková, D.; Poštová-Slavětínská, L.; Dračínský, M.;
Keough, D. T.; Guddat, L. W.; Janeba, Z. Acyclic Nucleoside Phosphonates
Containing 9-Deazahypoxanthine and a Five-Membered Heterocycle as Selective
Inhibitors of Plasmodial 6-Oxopurine Phosphoribosyltransferases. ChemMedChem
2017, 12, 1133–1141.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Špaček, P.; Keough, D. T.; Chavchich, M.; Dračínský, M.; Janeba, Z.; Naesens, L.;
Edstein, M. D.; Guddat, L. W.; Hocková, D. Synthesis and Evaluation of Asymmetric
Acyclic Nucleoside Bisphosphonates as Inhibitors of Plasmodium Falciparum and
Human Hypoxanthine–Guanine–(Xanthine) Phosphoribosyltransferase. J. Med. Chem.
2017, 60, 7539–7554.
Shah, R.; Verma, P. K. Therapeutic Importance of Synthetic Thiophene. Chemistry
Central Journal. BioMed Central December 19, 2018, p 137.
(10) Mishra, R.; Tomar, I.; Singhal, S.; Jha, K. K. Synthesis, Properties and Biological
Activity of Thiophene: A Review. Der Pharma Chem. 2011, 3, 38–54.
(11) Handbook of Thiophene-Based Materials; Perepichka, I. F., Perepichka, D. F., Eds.;
John Wiley & Sons, Ltd: Chichester, UK, 2009.
(12) Barbarella, G.; Zangoli, M.; Di Maria, F. Synthesis and Applications of Thiophene
Derivatives as Organic Materials. Adv. Heterocycl. Chem. 2017, 123, 105–167.
(13) Eicher, T.; Hauptmann, S.; Speicher, A. The Chemistry of Heterocycles; Wiley, 2003.
(14) Mancuso, R.; Gabriele, B. Recent Advances in the Synthesis of Thiophene Derivatives
by Cyclization of Functionalized Alkynes. Molecules. Multidisciplinary Digital
Publishing Institute September 29, 2014, pp 15687–15719.
(15) Haag, B.; Mosrin, M.; Ila, H.; Malakhov, V.; Knochel, P. Regio- and Chemoselective
Metalation of Arenes and Heteroarenes Using Hindered Metal Amide Bases. Angew.
Chemie - Int. Ed. 2011, 50, 9794–9824.
(16) Krasovskiy, A.; Krasovskaya, V.; Knochel, P. Mixed Mg/Li Amides of the Type
R2NMgCl.LiCl as Highly Efficient Bases for the Regioselective Generation of
25
ACS Paragon Plus Environment

The Journal of Organic Chemistry
Page 26 of 27
1
2
3
4
5
6
Functionalized Aryl and Heteroaryl Magnesium Compounds. Angew. Chemie - Int. Ed.
2006, 45, 2958–2961.
7
8
(17) Piller, F.; Knochel, P. Regio- and Chemoselective Synthesis of Fully Substituted
9
Furans. Synthesis (Stuttg). 2011, 2011, 1751–1758.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(18) Piller, F. M.; Knochel, P. Regio- and Chemoselective Synthesis of Fully Substituted
Thiophenes. Org. Lett. 2009, 11, 445–448.
(19) Ota Chikako; Kumata Shuji; Kawaguchi Shinji. Novel Herbicides, Usage Therof,
Novel Thienopyrimidine Derivatives, Intermediates of the Same, and Process for
Production Thereof. EP1544202 (A1), 2005.
(20) Beshore C. Douglas; Kuduk D. Scott; Liverton Nigel; Luo Yunfu; Meng Na; Yu
Tingting. Oxazole Orexin Receptor Antagonists. WO2015018029 (A1), 2015.
(21) Bloomfield, A. J.; Herzon, S. B. Room Temperature, Palladium-Mediated P-Arylation
of Secondary Phosphine Oxides. Org. Lett. 2012, 14, 4370–4373.
(22) Lennox, A. J. J.; Lloyd-Jones, G. C. Selection of Boron Reagents for Suzuki–Miyaura
Coupling. Chem. Soc. Rev. Chem. Soc. Rev 2014, 412, 412–443.
(23) Lozada, J.; Liu, Z.; Perrin, D. M. Base-Promoted Protodeboronation of 2,6-
Disubstituted Arylboronic Acids. J. Org. Chem. 2014, 79, 5365–5368.
(24) Chataigner, I.; Panel, C.; Gérard, H.; Piettre, S. R. Sulfonyl vs. Carbonyl Group: Which
Is the More Electron-Withdrawing? Chem. Commun. 2007, No. 31, 3288–3290.
(25) Krasovskiy, A.; Knochel, P. Eine LiCl-Vermittelte Br/Mg-Austauschreaktion Zur
Herstellung Funktionalisierter Aryl- Und Heteroarylmagnesium-Verbindungen
Ausgehend von Organischen Bromiden. Angew. Chemie 2004, 116, 3396–3399.
(26) Krasovskiy, A.; Krasovskaya, V.; Knochel, P. Gemischte Mg/Li-Amide Des Typs
R2NMgCl⋅LiCl Als Hoch Effiziente Basen Zur Regioselektiven Synthese
Funktionalisierter Aryl- Und Heteroarylmagnesium-Verbindungen. Angew. Chemie
2006, 118, 3024–3027.
(27) Monzon, G.; Knochel, P. Directed Ortho- and Meta-Magnesiation or Zincation of
Polyfunctional Aryl Nonaflates. Synlett 2010, 2010, 304–308.
(28) Melzig, L.; Rauhut, C. B.; Knochel, P. 2,3-Functionalization of Furans, Benzofurans
26
ACS Paragon Plus Environment

Page 27 of 27
The Journal of Organic Chemistry
1
2
3
4
5
6
and Thiophenes via Magnesiation and Sulfoxide-Magnesium Exchange. Chem.
Commun. 2009, No. 24, 3536–3538.
7
8
(29) Sawada, N.; Itoh, T.; Yasuda, N. Efficient Copper-Catalyzed Coupling of Aryl Iodides
9
and Thiobenzoic Acid. Tetrahedron Lett. 2006, 47, 6595–6597.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
(30) Wagner, R.; Donner, P. L.; Kempf, D. J.; Maring, C. J.; Stoll, V. S.; Ku, Y.; Pu, Y.
HCV Inhibitors. WO2008/11337, 2008.
27
ACS Paragon Plus Environment