Toll-like receptor-8 agonistic activities in C2, C4, and C8 modified thiazolo[4,5-c]quinolines

Article abstract of DOI:10.1039/c2ob26705e

Toll-like receptor (TLR)-8 agonists typified by the 2-alkylthiazolo[4,5-c] quinolin-4-amine (CL075) chemotype are uniquely potent in activating adaptive immune responses by inducing robust production of T helper 1-polarizing cytokines, suggesting that TLR8-active compounds could be promising candidate vaccine adjuvants, especially for neonatal vaccines. Alkylthiazoloquinolines with methyl, ethyl, propyl and butyl groups at C2 displayed comparable TLR8-agonistic potencies; activity diminished precipitously in the C2-pentyl compound, and higher homologues were inactive. The C2-butyl compound was unique in possessing substantial TLR7-agonistic activity. Analogues with branched alkyl groups at C2 displayed poor tolerance of terminal steric bulk. Virtually all modifications at C8 led to abrogation of agonistic activity. Alkylation on the C4-amine was not tolerated, whereas N-acyl analogues with short acyl groups (other than acetyl) retained TLR8 agonistic activity, but were substantially less water-soluble. Immunization in rabbits with a model subunit antigen adjuvanted with the lead C2-butyl thiazoloquinoline showed enhancements of antigen-specific antibody titers.

Full text of DOI:10.1039/c2ob26705e

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Toll-like receptor-8 agonistic activities in C2, C4, and
C8 modified thiazolo[4,5-c]quinolines†
Cite this: Org. Biomol. Chem., 2013, 11,
1179
Hari Prasad Kokatla, Euna Yoo, Deepak B. Salunke, Diptesh Sil, Cameron F. Ng,
Rajalakshmi Balakrishna, Subbalakshmi S. Malladi, Lauren M. Fox and Sunil A. David*
Toll-like receptor (TLR)-8 agonists typified by the 2-alkylthiazolo[4,5-c]quinolin-4-amine (CL075) chemo-
type are uniquely potent in activating adaptive immune responses by inducing robust production of
T helper 1-polarizing cytokines, suggesting that TLR8-active compounds could be promising candidate
vaccine adjuvants, especially for neonatal vaccines. Alkylthiazoloquinolines with methyl, ethyl, propyl and
butyl groups at C2 displayed comparable TLR8-agonistic potencies; activity diminished precipitously in
the C2-pentyl compound, and higher homologues were inactive. The C2-butyl compound was unique in
possessing substantial TLR7-agonistic activity. Analogues with branched alkyl groups at C2 displayed
poor tolerance of terminal steric bulk. Virtually all modifications at C8 led to abrogation of agonistic
activity. Alkylation on the C4-amine was not tolerated, whereas N-acyl analogues with short acyl groups
(other than acetyl) retained TLR8 agonistic activity, but were substantially less water-soluble. Immuniz-
ation in rabbits with a model subunit antigen adjuvanted with the lead C2-butyl thiazoloquinoline
showed enhancements of antigen-specific antibody titers.
Received 30th August 2012,
Accepted 21st December 2012
DOI: 10.1039/c2ob26705e
www.rsc.org/obc
encoded in the human genome, which are trans-membrane
proteins with an extracellular domain having leucine-rich
Introduction
There can be no greater substantiation of Benjamin Franklin’s repeats (LRR) and a cytosolic domain called the Toll/IL-1 recep-
adage, ‘An ounce of prevention is better than a cure’ than the tor (TIR) domain.6 The ligands for these receptors are highly
resounding impact that vaccines have had in preventing mor- conserved molecules such as lipopolysaccharides (LPS) (recog-
bidity and mortality due to infectious diseases.1 Vaccines nized by TLR4), lipopeptides (TLR2 in combination with TLR1
afford protection by the induction of immune responses, both or TLR6), flagellin (TLR5), single stranded RNA (TLR7 and
humoral and cellular, specifically directed against the patho- TLR8), double stranded RNA (TLR3), CpG motif-containing
gen. Such adaptive immune responses are mobilized and DNA (recognized by TLR9), and profilin present on uropatho-
amplified by engagement of the innate immune system, a phy- genic bacteria (TLR11).10 TLR1, -2, -4, -5, and -6 recognize
logenetically and teleologically ancient system of host extracellular stimuli, while TLR3, -7, -8 and -9 function within
defense.2–4 Unlike adaptive immunity, the initial innate the endolysosomal compartment.6
immune responses rely on a limited number of germline-
The activation of TLRs by their cognate ligands leads to pro-
encoded pattern recognition receptors which recognize duction of inflammatory cytokines, and up-regulation of MHC
specific molecular patterns present in molecules that are molecules and co-stimulatory signals in antigen-presenting
broadly shared by pathogens but are sufficiently different so as cells as well as activating natural killer (NK) cells (innate
to be distinguishable from host molecules.5,6
immune response), which leads to the priming and amplifica-
Innate immune afferent signals include those originating tion of T-, and B-cell effector functions (adaptive immune
from Toll-like receptors (TLRs), as well as RIG-I-like receptors7 responses).11–14
and Nod-like receptors (NLRs).8,9 There are 10 functional TLRs
In recognition that the initial innate immune responses
serve to marshal, focus, and amplify subsequent adaptive
immune responses,15,16 there is considerable interest in utiliz-
ing TLR agonists as vaccine adjuvants.17,18 In contradistinction
to early vaccines which used killed whole organisms,19–22 or
attenuated live vaccines,23–25 modern vaccines increasingly rely
on subunit vaccines which have the distinct advantages of ease
of production, quality control, and safety; however, such
Department of Medicinal Chemistry, University of Kansas, Multidisciplinary
Research Building, Room 320D, 2030 Becker Drive, Lawrence, KS 66047, USA.
E-mail: sdavid@ku.edu; Fax: +1-785-864-1961; Tel: +1-785-864-1610
†Electronic supplementary information (ESI) available: Characterization of inter-
mediates and final compounds (1H, 13C, mass spectra). See DOI: 10.1039/
c2ob26705e
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subunit antigens which are largely soluble proteins are poorly
immunogenic, and require adjuvants to induce robust
immune responses. Currently, the only TLR agonist approved
by the FDA as an adjuvant is 3-O-desacyl-4′-monophosphoryl
lipid A, (MPL), a TLR4 agonist derived from hydrolytic treat-
ment of lipopolysaccharide isolated from Salmonella minnesota
Re595.26
We have recently begun exploring27,28 a variety of TLR ago-
nists with a view to identifying safe and potent vaccine adju-
vants. The chemotypes that we have explored thus far include
agonists of TLR2,29–31 TLR7,32–36 nucleotide oligomerization
domain 1 (Nod1),37 as well as C–C chemokine receptor type 1
(CCR1).38 We now extend our investigations toward delineating
structure–activity relationships in small-molecule agonistic
ligands of TLR8 (typified by the thiazolo[4,5-c]quinolines) for
the following reasons:
First, the thiazoloquinolines are of interest because like the
imidazoquinolines, these compounds were identified in anti-
Scheme 1 Syntheses of C2-alkylthiazoloquinoline analogues. Reagents: (i) HCl,
HONvCHCH2NO2, H2O; (ii) (CH3CO)2O, CH3COOK; (iii) Pt/C, H2, DMF; (iv) RCOCl,
viral assays long before the discovery of the endosomal TLR7 Et3N, CH2Cl2 : DMF(10 : 1); (v) P2S5, pyridine; (vi) m-CPBA, CHCl3; (vii) (a) benzoyl
and TLR8 receptors,39–43 and other than the original landmark
isocyanate, CH2Cl2, (b) NaOCH3, MeOH.
studies performed by investigators at 3M Pharmaceuticals,44,45
structure–activity relationships in the thiazoloquinoline chemo-
Given the potential utility of TLR8 agonists as adjuvants for
neonatal vaccines, it was of interest to explore SAR in the thi-
azoloquinolines (typified by CL075, 8c in Scheme 1). Our
studies began with examining the optimal alkyl chain length
at the C2 position. We found that the C2-alkyl thiazoloquino-
lines exhibit mixed TLR8/7 agonistic activities in primary
reporter gene assays with the optimal chain length being
butyl. We observed an unexpectedly strict length dependence
with only the C2-butyl, but none of the other analogues, indu-
cing IFN-α in human peripheral blood mononuclear cells
(hPBMCs). Examination of analogues with branched alkyl
groups at C2 suggested poor tolerance of terminal steric bulk.
We noted, however that certain C2-branched analogues were
substantially more TLR8-selective than their corresponding
straight-chain analogues. Virtually all modifications at C8 led
to abrogation of agonistic activity. Alkylation on the C4-amine
was not tolerated, whereas N-acyl analogues with short acyl
groups (other than acetyl) retained activity.
type remains poorly explored; qualitative assays for TNF-α
and IFN-α induction in human blood were performed in initial
studies by 3M as surrogate biomarkers of immunostimulation,
and no data on TLR-7 and -8 specific agonistic activities exist
in the literature.
Second, TLR8 agonists, both single-stranded RNA as well as
small molecule imidazoquinoline ligands such as R-84846 and
thiazoloquinolines such as 3M-002 (CL075)47,48 appear
uniquely potent in activating co-stimulatory responses in neo-
natal antigen-presenting cells (APCs), inducing robust pro-
duction of the T helper 1 (Th1)-polarizing cytokines TNF-α and
IL-12, which are not observed upon stimulation by TLR-2, -4,
or -7 agonists. Such Th1-biasing compounds are of particular
interest as candidate vaccine adjuvants in the newborn.49
During the first few weeks of life, newborns rely almost entirely
on maternal IgG antibodies acquired by passive transplacental
passage,50 and remain susceptible to a wide range of patho-
gens until early infancy.51 Neonates and infants, in whom vac-
cines could—and perhaps should—have the greatest impact
do not mount adequate adaptive immune responses, and
therefore are most vulnerable; consequently, even the most
efficacious vaccines that confer excellent protection in adults
Results and discussion
may fail to elicit strong immune responses in them.52,53 There In our previous work on the TLR7-active imidazoquinolines,
is mounting evidence pointing to significant differences we had observed a distinct relationship between C2-alkyl chain
between adult and infant innate immune responses.54 The length and TLR7-agonistic potency,33 and we therefore
neonatal immunophenotype is characterized by decreased pro- thought it logical to begin our SAR studies on the thiazoloqui-
duction of both type I and type II interferons, Interleukin (IL)- nolines by examining analogues with C2-alkyl groups of
12, IL-18, IL-23 and other proinflammatory cytokines such as varying chain lengths. These analogues (8a–8h, Scheme 1)
Tumor Necrosis Factor-α (TNF-α), the preferential induction of were synthesized in parallel from the 3-aminoquinolin-4-ol
memory B lymphocytes rather than immunoglobulin-secreting precursor 4, using the strategy described earlier.33,44
plasma cells, as well as a pronounced T-helper type 2 (Th2)
In primary screens using human TLR7 and TLR8-specific
skewing of T-cell responses.51,52,55,56 TLR8 agonists induce the reporter gene assays, these analogues exhibited mixed TLR8/
production of IL-12, IL-18 and IFN-γ, and may therefore be of TLR7 agonism; the EC50 values of 8c (CL075) were found to be
value in developing vaccines for the neonate.
1.32 μM and 5.48 μM, in TLR8 and TLR7 assays, respectively
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parent compounds (Fig. 3), nor did they show enhanced IL-12,
IL-18, and IFN-γ production (data not shown) as would be
expected for TLR8-selective compounds.47,48 We do not yet
understand the basis of attenuated activity of the C2-branched
analogues, and we are currently exploring whether differential
plasma protein binding behavior of these compounds could
be contributory.57
Previous reports on the thiazoloquinolines44,45,58 had been
performed largely on the C7 position, and C8 and C4
analogues
remain
unexplored.
We
therefore
first
examined substituents on the quinoline at the C8 position of
8c (CL075). Electrophilic substitution on the quinoline
ring selectively afforded the 8-nitro analogue 9 (Scheme 2),
and the 8-bromo analogue 14 (Scheme 4), both of which
were inactive. The 8-amino (10) and 8-azido (11) analogues
were obtained from 9 (Scheme 2). Compound 10 showed
attenuated activity relative to 8c, and 11 was inactive. The
triazole derivatives 12a–g were synthesized from 11 (Scheme 2)
using conventional copper-catalyzed ‘click’ chemistry; of
Fig. 1 TLR8 and TLR7 agonistic potencies of the C2-alkyl thiazoloquinoline
homologues. Data points represent means and standard deviations of EC50
values derived from dose-response profiles and are computed on
quadruplicates.
(Fig. 1, Table 1). As in our earlier SAR studies on the imidazo- these analogues, the triazolo analogue 12e displayed feeble,
quinolines,33 we observed a clear dependence of agonistic
but selective TLR8 agonism (Table 1). Selective C8 N-alkylation
potency on the C2-alkyl chain length. The C2-methyl, -ethyl,
and -propyl analogues (8a–8c) displayed comparable potencies logues 13a and 13b (Scheme 3), both of which were inactive
and N-acylation of the 8-amino analogue 10 provided ana-
in the hTLR8 and hTLR7 reporter gene assays (Fig. 1).
Maximal TLR8-agonistic potency was observed in the butyl
(Table 1).
Next, the C4-amine of 8c was alkylated or acylated, yielding
analogue (8d). Increasing the chain length to pentyl (8e) led to analogues 15a–15l (Scheme 5). The C4-N-alkylated compounds
attenuation of potency, and higher homologues (8f–8h)
showed abrogation of activity (Fig. 1, Table 1).
15a and 15b were devoid of any agonistic activity, while some,
but not all of the C4-N-acylated derivatives with short acyl
The C2-butyl analogue 8d appeared to exhibit substantially groups were found to be active. Specifically, the formyl (15d),
higher TLR7-agonistic potency than 8c (Fig.
1 and 3),
and butyryl (15f) analogues, but not the acetyl (15e) com-
pounds were active.
Although weaker in potency than the parent compound 8c,
these C4-N-acyl derivatives exhibited a pronounced specificity
for TLR8 (Table 1). We observed an unusual discontinuity in
suggesting that small variations at C2 could modulate TLR8
versus TLR7 specificity. The introduction of an isopropyl group
at C2 (8i) led to an approximately ten-fold reduction in TLR8-
agonistic potency (Table 1), while the attenuation of its activity
in TLR7-specific reporter gene assays was more modest (about the activity profiles in this homologous series of compounds.
two-fold). These results warranted the syntheses of several
additional analogues of both 8c (C2-n-propyl) and 8d (C2-n-
The TLR8-agonistic potency of the formyl analogue 15d was
comparable to that of 8c. The acetyl analogue 15e, however,
butyl) with branched alkyl groups at C2 (8j–8o, Scheme 1, was virtually inactive, but activity was partially restored in the
Fig. 2). Additional methyl substitutions at ω − 1 as well as ω − 2
positions were well tolerated in 8c derivatives (8j and 8n,
respectively; Scheme 1, Fig. 2). On the other hand only the ω − 2
methyl-substituted compound 8o was active in the 8d series,
whereas the 8l congener with a terminal methyl substitution
was entirely inactive. Both 8k and 8m bearing terminal tri-
methyl groups were also inactive, suggesting poor tolerance of
terminal steric bulk in these compounds.
butyryl compound 15f (Fig. 4A), while higher homologues (15g
and 15h) were inactive (Table 1).
Cytokine induction profiles in hPBMCs mirrored these find-
ings with a near-complete loss of activity for the acetyl com-
pound 15d (Fig. 5A).
Our provisional interpretation that a strict length require-
ment exists for the acyl substituents on the C4-amine was
borne out in compounds 15j–15l (azidoacetamide, azidopro-
We also evaluated bioisosteric analogues 8p and 8q pionamide and pentynamide analogues, respectively) which
(Scheme 1) with terminal trifluoromethyl groups. A decrease
in potency at TLR8 was observed in the longer homologue
displayed weak, TLR8-selective agonistic activity (Table 1). Aro-
matic amides at this position appear not to be tolerated, since
(8q). We noted that, in general, analogues with branched C2- the benzamide analogue 15i was entirely inactive.
alkyl groups of optimal chain length and bulk (such as 8j, 8n,
and 8o) are substantially more TLR8-selective than their corre-
sponding straight-chain analogues (Fig. 2).
However, in secondary cytokine induction screens using
human PBMCs, the branched-chain analogues behave differ-
A series of carbamates (15m–15p), and sulfonamides (15r–
15v) were also synthesized (Scheme 5). The carbamate deriva-
tives showed very weak activity in primary reporter gene assays,
while the sulfonamide analogues were completely inactive
(Table 1). Several other analogs including urea (15q;
ently in that they were neither as potent as their straight-chain Scheme 5), phosphoramidate (15w; Scheme 5) and guanidine
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Table 1 EC50 values of thiazoloquinolines in human TLR8- and TLR7-specific
reporter gene assays
Table 1 (Contd.)
Agonistic
activity (μM)
Agonistic
activity (μM)
Compound
number
Compound
R1
R2
R3
TLR8
TLR7
number
R1
R2
H
R3
TLR8
9.84
TLR7
8a
8b
8c
8d
8e
8f
8g
8h
8i
CH3
C2H5
C3H7
C4H9
C5H11
C6H13
C7H15
C9H19
CH(CH3)2
CH2CH(CH3)2
CH2C(CH3)3
C2H5CH
(CH3)2
C2H5C(CH3)3
(S)-2-(sec-
Butyl)
(S)-2-(2-
Methylbutyl)
C2H5CF3
C3H7CF3
C3H7
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
2.06
0.81
1.32
0.41
1.94
10.24
5.12
5.48
0.86
4.14
12f
C3H7
Inactive
12g
C3H7
H
Inactive Inactive
Inactive Inactive
Inactivea Inactive
Inactive Inactive
Inactive Inactive
13a
13b
14
15a
15b
15c
15d
15e
15f
15g
15h
15i
15j
15k
15l
15m
15n
15o
15p
15q
15r
15s
15t
15u
15v
15w
17
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C3H7
C4H9
C4H9
C4H9
H
H
H
C4H9
C6H13
C16H33
CHO
COCH3
COC3H7
COC7H15
COC15H31
COC5H5
COCH2N3
COC2H4N3
C6H13
10.85
1.55
Inactive Inactive
Inactive Inactive
9.67
6.96
COC3H7 Inactive Inactive
8j
8k
8l
Br
H
H
H
H
H
H
H
H
H
H
H
Inactive Inactive
Inactive Inactive
Inactive Inactive
Inactive Inactive
Inactive Inactive
0.88
1.14
Inactive Inactive
Inactive Inactive
Inactive Inactive
1.20
2.23
1.20
Weak
8m
8n
H
H
H
H
Inactive Inactive
2.77
8.11
Inactive
Inactive
8o
H
H
2.51
4.74
8p
8q
9
10
11
H
H
H
H
H
H
H
NO2
NH2
N3
1.61
6.29
3.63
2.25
3.53
Inactive Inactive
3.88
2.06
Inactive
Inactive
Inactive
C3H7
C3H7
8.19
10.24
COC2H4CuC H
CO2CH3
CO2C2H5
CO2C4H9
CO2C8H17
CONH2
SO2CH3
SO2C2H5
SO2C3H7
SO2C4H9
SO2C6H4CH3
PO(OEt)2
CNHNH2
CHO
H
H
H
H
H
H
H
H
H
H
H
H
H
H
H
12a
C3H7
H
Inactive Inactive
Inactive Inactive
Inactive Inactive
Inactive Inactive
Inactive Inactive
10.56
Inactive
Inactive Inactive
Inactive Inactive
Inactive Inactive
Inactive Inactive
Inactive Inactive
Inactive Inactive
2.31
Inactive Inactive
0.42 0.49
12b
C3H7
H
Inactive Inactive
12c
12d
C3H7
C3H7
H
H
Inactive Inactive
Inactive Inactive
18a
18b
18c
2.54
COCH3
COC3H7
a EC50 value is >100 μM.
(18a–18c, formamide, acetamide, and butyramide derivatives,
respectively; Scheme 7). As with the 8c derivatives, a virtually
identical pattern was observed, with a selective loss of activity
for the acetamide analogue 18b (Fig. 4B, 5B, Table 1).
12e
C3H7
H
Inactive Inactive
Our studies aimed at delinating structure–activity relation-
ships in the thiazoloquinolines have led to selecting 8d as a
lead thiazoloquinoline. This compound distinguishes itself
from its closely-related congener, 8c (CL075) in possessing a
more balanced TLR8/TLR7-agonistic properties. Virtually all
other analogues were either of lower potency, exhibited very
poor aqueous solubility (such as the amide analogues 15d and
18a), or despite apparent greater TLR8 selectivity in primary
(17; Scheme 6) functional groups at C4 were also examined,
but were found to be inactive.
The peculiar SAR, with only the acetamide analogue (15e)
showing loss of activity was unexpected and, desiring to confirm
this pattern, we also synthesized select amide analogues of 8d
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Scheme 2 Modification at the C8 position. Reagents: (i) HNO3, H2SO4; (ii) Zn,
NH4COOH, MeOH; (iii) NaNO2, CH3COOH, NaN3; (iv) alkyne, CuSO4, sodium
ascorbate, THF, H2O (v) TBAF, THF.
Fig. 2 TLR8- and TLR7-specific NF-κB induction profiles of branched-chain and
trifluoromethyl analogues of 8c and 8d. Means and SD obtained from quadru-
plicate samples are shown.
Scheme 3 Synthesis of C8 N-alkyl and N-acyl analogues. Reagents: (i) For 13a,
C6H13I, K2CO3, DMF; For 13b, C3H7COCl, Et3N, CH2Cl2.
Scheme 4 Synthesis of 8-bromothiazoloquinoline. Reagents: (i) NBS, NH4OAc,
CH3CN.
screens, were either unremarkable in secondary screens
employing human PBMCs, or inferior in their cytokine induc-
tion profiles (8j, 8n, and 8o, for instance). We therefore elected
to directly evaluate the adjuvantic properties of 8d, and bench-
mark it against 8c (CL075), noting that adjuvantic properties
for the thiazoloquinolines have not been previously reported
in the literature. In light of the fact that murine TLR8 is
thought to be functionally inactive,59–61 we chose to examine
the adjuvantic activities of 8c, 8d directly in a rabbit model
using bovine α-lactalbumin as a model subunit vaccine
Fig. 3 EC50 values of proinflammatory cytokine induction in human PBMCs by
analogues of 8c and 8d. Representative data from three independent experi-
ments are shown.
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Scheme 6 Syntheses of C4 guanidino analog. Reagents: (i) POCl3, 100 °C; (ii)
guanidine hydrochloride, NaH, 1,4-dioxane.
Scheme 5 Syntheses of C4 N-alkyl and N-acyl analogues. Reagents: (i) for 15a,
15b and 15c, RX, NaH, THF; for 15d, 2,2,2-trifluoroethylformate, Et3N; for 15e–i,
RCOCl, Py; for 15j–l, RCO2H, HBTU, Et3N; for 15m–p, ROCOCl, Et3N, CH2Cl2; for
15q, ClSO2NCO, NaHCO3; for 15r–v, RSO2Cl, CH2Cl2; for 15w, ClP(O)OEt2,
CH2Cl2.
Scheme 7 Syntheses of C4 N-alkyl and N-acyl analogues of 8d. Reagents: (i)
2,2,2-trifluoroethylformate, Et3N; For 18a; RCOCl, pyridine for 18b and 18c.
Fig. 4 TLR8 induction by C4-amides of 8c (Panel A) and 8d (Panel B). Data
points represent means and standard deviations on quadruplicates.
Fig. 6 Box-plots of anti-bovine α-lactalbumin IgG titers in cohorts of three
rabbits immunized with α-lactalbumin adjuvanted with either 8c or 8d. Means
□
and medians of titers are represented by
and ─ symbols within the box,
respectively, and the × symbols indicate the 1% and 99% percentile values.
antigen, which is a small (14 kDa), soluble protein which we
have adopted as our test-antigen in ongoing humoral and cel-
lular immune response assays.35 We found that both 8c and
8d were adjuvantic in evoking high antigen-specific IgG titers
(Fig. 6). Importantly, no evidence of local or systemic toxicity
was apparent in any of the cohorts. The data suggests that 8d,
with its dual TLR8/TLR7-agonistic properties, elicits adjuvanti-
city with greater consistency and uniformity, as evidenced by
narrower confidence intervals of antibody titers (Fig. 6).
Fig. 5 Dose–response profiles of proinflammatory cytokine induction in
hPBMCs by C4-amides of 8c (Panel A) and 8d (Panel B). Representative data
from three independent experiments are presented. Vehicle controls (not
shown) elicited undetectable levels of cytokines.
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2 (12.94 g, 85%). 1H NMR (400 MHz, MeOD) δ 8.14 (d, J =
7.7 Hz, 1H), 7.83 (d, J = 6.1 Hz, 1H), 7.68–7.59 (m, 2H), 7.22
Conclusions
TLR8 agonists are thought to be uniquely potent in activating (t, J = 7.4 Hz, 1H), 6.73 (d, J = 6.3 Hz, 1H). 13C NMR (101 MHz,
adaptive immune responses by inducing robust production MeOD) δ 168.6, 141.2, 136.6, 134.3, 132.0, 123.2, 116.7, 114.5,
of T helper 1-polarizing cytokines, and may be promising can- 100.0. MS (ESI) calculated for C9H8N2O4, m/z 208.05, found
didate vaccine adjuvants, especially for neonatal vaccines. 209.06 (M + H)+.
Thiazoloquinoline analogues with methyl, ethyl, propyl and
butyl groups at C2 displayed comparable TLR8-agonistic Synthesis of compound 3: 3-nitroquinolin-4-ol
potencies; however, the C2-butyl compound 8d was unique in
A solution of compound 2 (12.94 g, 62.2 mmol) in acetic anhy-
possessing substantial TLR7-agonistic activity. Analogues with
dride (50 mL) was placed in a 2-neck flask fitted with a reflux
branched alkyl groups at C2 displayed poor tolerance of term-
condenser. It was stirred and heated to 105 °C until a clear solu-
inal steric bulk. C4-N-acyl analogues with short acyl groups
tion was obtained. Heating was then discontinued and pot-
(other than acetyl) retained TLR8 agonistic activity, but were
assium acetate (6.22 g, 63.5 mmol) was added. The mixture
substantially less water-soluble. Immunization in rabbits with
was then refluxed for 15 min with vigorous stirring, until a
a model subunit antigen adjuvanted with the most potent
solid started to precipitate. The reaction mixture was then
TLR8 agonist showed dramatic enhancements of antigen-
slowly cooled to room temperature. The residue was filtered,
specific antibody titers.
washed with glacial acetic acid until the washings were color-
less, then suspended in water, filtered, washed with water and
dried at 110 °C to get 3-nitroquinolin-4-ol 3 (4.68 g, 40%).
1H NMR (500 MHz, DMSO) δ 13.04 (s, 1H), 9.21 (s, 1H), 8.25
(dd, J = 8.1, 1.1 Hz, 1H), 7.83–7.77 (m, 1H), 7.74–7.70 (m, 1H),
7.52 (ddd, J = 8.1, 7.1, 1.1 Hz, 1H). 13C NMR (126 MHz, DMSO)
δ 167.6, 142.5, 138.3, 133.2, 130.9, 128.1, 125.9, 125.8, 119.5.
MS (ESI) calculated for C9H6N2O3, m/z 190.04, found 191.05
(M + H)+.
Experimental
Materials and equipment
All of the solvents and reagents used were obtained commer-
cially and used as such unless noted otherwise. Moisture- or
air-sensitive reactions were conducted under nitrogen atmos-
phere in oven-dried (120 °C) glass apparatus. The solvents
were removed under reduced pressure using standard rotary
evaporators. Flash column chromatography was carried out
using RediSep Rf ‘Gold’ high performance silica columns on
CombiFlash Rf instrument unless otherwise mentioned, while
thin-layer chromatography was carried out on silica gel
(200 μm) CCM pre-coated aluminum sheets. Purity for all final
compounds was confirmed to be greater than 98% by LC-MS
using a Zorbax Eclipse Plus 4.6 mm × 150 mm, 5 μm analytical
reverse phase C18 column with H2O–isopropanol or H2O–
CH3CN gradients and either an Agilent ESI-TOF mass spec-
trometer (mass accuracy of 20 ppm) or an Agilent 6520
ESI-QTOF mass spectrometer (mass accuracy of <10 ppm)
operating in the positive ion (or negative ion, as appropriate)
acquisition mode.
Synthesis of compound 4: 3-aminoquinolin-4-ol
To a solution of compound 3 (1.89 g, 9.93 mmol) in DMF
(25 mL), was added 5% Pt on carbon (20%, 0.38 g). The reac-
tion mixture was allowed to react in a Parr hydrogenation
apparatus at 60 psi H2 pressure for 3.5 h with vigorous
shaking. The reaction mixture was filtered through celite with
several washes of methanol. The filtrate was concentrated by
evaporation to get 4-amino-3-nitroquinoline (1.5 g, 94%).
1H NMR (500 MHz, MeOD) δ 8.90 (d, J = 8.1 Hz, 1H), 8.33
(d, J = 5.8 Hz, 1H), 8.30–8.23 (m, 2H), 7.97 (ddd, J = 8.1, 6.0,
1.9 Hz, 1H). 13C NMR (126 MHz, MeOD) δ 146.9, 140.8, 138.9,
134.2, 131.2, 130.6, 128.7, 127.5. MS (ESI) calculated for
C9H8N2O, m/z 160.06, found 161.07 (M + H)+.
Synthesis of compound 2: 2-(2-nitroethylideneamino)benzoic
acid
Synthesis of compound 5a: N-(4-hydroxyquinolin-3-yl)-
acetamide
Nitromethane (4.32 mL, 80 mmol) was added dropwise to a Compound 4 (200 mg, 1.25 mmol) was dissolved in a mixture
solution of NaOH (9.6 g, 250 mmol) in water (10 mL), at 0 °C. of CH2Cl2 (20 mL) and DMF (2 mL) and stirred at room temp-
The mixture was then warmed to 40 °C and nitromethane erature for 5 min. Acetyl chloride (133 μL, 1.875 mmol) was
(4.32 mL, 80 mmol) was again added slowly at 40–45 °C. The added to the stirring reaction mixture at 0 °C and the solution
temperature was maintained until
a clear solution was was allowed to react for 1.5 h. Solvents were removed and the
obtained. The reaction mixture was then heated to 55 °C for crude residue was purified using silica gel column chromato-
2–5 minutes, cooled to 30 °C, poured onto crushed ice and graphy (10% MeOH/CH2Cl2) to obtain the compound 5a
acidified with conc. HCl (11 mL). The resultant solution of (106 mg, 42%). 1H NMR (500 MHz, CDCl3) δ 9.20 (d, J = 5.5 Hz,
methazoic acid was added immediately to a filtered solution of 1H), 8.86 (s, 1H), 8.46 (s, 1H), 8.41 (dd, J = 8.3, 1.0 Hz, 1H),
anthranilic acid 1 (10 g, 73 mmol) and conc. HCl (3.3 mL) in 7.62 (ddd, J = 8.3, 7.1, 1.3 Hz, 1H), 7.41–7.32 (m, 2H), 2.26
water (75 mL). The reaction mixture was allowed to stand at (s, 3H). 13C NMR (126 MHz, CDCl3)δ 169.1, 137.9, 132.1, 126.6,
room temperature for 12 h. After filtration, the residue 126.3, 123.6, 123.3, 122.9, 117.6, 24.6. MS (ESI) calculated for
obtained was washed with water, and dried to yield compound C11H10N2O2, m/z 202.07, found 203.08 (M + H)+.
This journal is © The Royal Society of Chemistry 2013
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Compounds 5b–5q were synthesized similarly as compound 22.8, 14.2. MS (ESI) calculated for C17H22N2O2, m/z 286.17,
5a.
found 287.18 (M + H)+.
5b: N-(4-Hydroxyquinolin-3-yl)propionamide
5h: N-(4-Hydroxyquinolin-3-yl)decanamide
1
1
(132 mg, 43%). H NMR (500 MHz, CDCl3) δ 9.29 (s, 1H), 8.71 (231 mg, 59%). H NMR (500 MHz, CDCl3) δ 9.24 (s, 1H), 8.47
(s, 1H), 8.42 (d, J = 7.7 Hz, 1H), 7.64 (t, J = 7.4 Hz, 1H), 7.49 (s, (s, 1H), 8.41 (dd, J = 8.2, 1.0 Hz, 1H), 7.63–7.58 (m, 1H), 7.40
1H), 7.38 (s, 1H), 2.56 (dd, J = 14.2, 7.0 Hz, 2H), 1.31 (t, J = 7.5 (d, J = 8.4 Hz, 1H), 7.37–7.32 (m, 1H), 2.50–2.43 (m, 2H),
Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 137.9, 132.2, 126.1, 1.80–1.72 (m, 2H), 1.43–1.20 (m, 12H), 0.87 (t, J = 7.0 Hz, 3H).
126.1, 124.5, 124.1, 124.0, 123.9, 118.0, 30.6, 9.9. MS (ESI) cal- 13C NMR (126 MHz, CDCl3) δ 172.4, 170.8, 138.0, 132.0, 126.8,
culated for C12H12N2O2, m/z 216.09, found 217.10 (M + H)+.
126.2, 123.5, 123.3, 122.7, 117.7, 37.7, 32.0, 29.6, 29.5, 29.4,
29.4, 25.9, 22.8, 14.3. MS (ESI) calculated for C19H26N2O2, m/z
314.20, found 315.21 (M + H)+.
5c: N-(4-Hydroxyquinolin-3-yl)butyramide
1H NMR (500 MHz, MeOD) δ 9.02 (s, 1H), 8.28 (dd, J = 8.3, 1.3
Hz, 1H), 7.68 (ddd, J = 8.4, 7.0, 1.4 Hz, 1H), 7.57 (d, J = 8.4 Hz,
Synthesis of compound 6a: 2-methylthiazolo[4,5-c]quinoline
1H), 7.42–7.35 (m, 1H), 2.46 (t, J = 7.5 Hz, 2H), 1.74 (dd, J = To a solution of compound 5a (100 mg, 0.49 mmol) in pyridine
14.9, 7.4 Hz, 2H), 1.01 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, (10 mL), was added phosphorous pentasulfide (109 mg,
MeOD) δ 174.6, 172.2, 139.7, 133.1, 131.3, 126.2, 124.8, 124.7, 0.49 mmol) and reaction mixture was refluxed for 2 h. The
122.8, 119.4, 39.6, 20.3, 14.0. MS (ESI) calculated for resulting solution was cooled to room temperature and pyri-
C13H14N2O2, m/z 230.11, found 231.11 (M + H)+.
dine was removed under reduced pressure. The residue was
dissolved in water and pH was adjusted to 8 with saturated
sodium bicarbonate solution and extracted in ethyl acetate.
5d: N-(4-Hydroxyquinolin-3-yl)pentanamide
(155 mg, 52%). 1H NMR (500 MHz, CDCl3) δ 10.61 (s, 1H), 9.18 The organic layer was dried over sodium sulfate and concen-
(s, 1H), 8.98 (s, 1H), 8.19 (d, J = 8.0 Hz, 1H), 7.72–7.62 (m, 2H), trated to afford compound 6a (64 mg, 65%). 1H NMR
7.37–7.30 (m, 1H), 2.44 (t, J = 7.4 Hz, 2H), 1.60–1.52 (m, 2H), (500 MHz, CDCl3) δ 9.42 (s, 1H), 8.23 (d, J = 8.3 Hz, 1H), 7.94
1.32 (dq, J = 14.7, 7.4 Hz, 2H), 0.89 (dd, J = 12.5, 5.1 Hz, 3H). (dd, J = 8.1, 0.9 Hz, 1H), 7.73 (ddd, J = 8.4, 7.0, 1.4 Hz, 1H),
13C NMR (126 MHz, CDCl3) δ 171.6, 168.9, 138.0, 131.4, 129.1, 7.62 (ddd, J = 8.1, 7.1, 1.1 Hz, 1H), 2.94 (s, 3H). 13C NMR
124.0, 123.0, 122.9, 121.4, 118.5, 27.4, 21.9, 13.8. MS (ESI) cal- (126 MHz, CDCl3) δ 167.5, 148.0, 145.6, 144.3, 141.1, 130.6,
culated for C14H16N2O2, m/z 244.12, found 245.13 (M + H)+.
128.8, 127.6, 125.0, 123.5, 20.3. MS (ESI) calculated for
C11H8N2S, m/z 200.04, found 201.05 (M + H)+.
5e: N-(4-Hydroxyquinolin-3-yl)hexanamide
Compounds 6b–6q were synthesized similarly as compound
1
(141 mg, 44%). H NMR (500 MHz, CDCl3) δ 9.65 (s, 1H), 8.44 6a.
(d, J = 8.2 Hz, 1H), 8.31 (d, J = 7.2 Hz, 1H), 7.88 (t, J = 7.4 Hz,
1H), 7.68 (t, J = 7.3 Hz, 1H), 2.82 (t, J = 6.5 Hz, 2H), 1.88–1.76
6b: 2-Ethylthiazolo[4,5-c]quinoline
1
(m, 2H), 1.48–1.35 (m, 4H), 0.92 (t, J = 7.0 Hz, 3H). 13C NMR (90 mg, 70%). H NMR (500 MHz, CDCl3) δ 9.45 (s, 1H), 8.24
(126 MHz, CDCl3) δ 177.9, 137.1, 133.7, 128.0, 124.8, 122.0, (d, J = 8.3 Hz, 1H), 7.99–7.94 (m, 1H), 7.73 (ddd, J = 8.4, 7.0,
120.7, 120.1, 36.1, 31.3, 25.6, 22.5, 14.1. MS (ESI) calculated for 1.4 Hz, 1H), 7.64 (ddd, J = 8.1, 7.1, 1.1 Hz, 1H), 3.26 (q, J = 7.6
C15H18N2O2, m/z 258.14, found 259.15 (M + H)+.
Hz, 2H), 1.54 (dd, J = 10.6, 4.5 Hz, 3H). 13C NMR (126 MHz,
CDCl3) δ 174.2, 147.9, 145.8, 144.2, 140.6, 130.6, 128.8, 127.6,
125.0, 123.6, 28.0, 14.0. MS (ESI) calculated for C12H10N2S, m/z
5f: N-(4-Hydroxyquinolin-3-yl)heptanamide
(105 mg, 31%). 1H NMR (500 MHz, CDCl3) δ 9.24 (d, J = 6.3 Hz, 214.06, found 215.08 (M + H)+.
1H), 8.88 (d, J = 1.6 Hz, 1H), 8.46 (s, 1H), 8.43–8.40 (m, 1H),
7.62 (ddd, J = 8.4, 7.1, 1.4 Hz, 1H), 7.40–7.33 (m, 2H),
2.49–2.44 (m, 2H), 1.80–1.72 (m, 2H), 1.44–1.36 (m, 2H), 1.32 (950 mg, 95%). H NMR (400 MHz, CDCl3) δ 9.45 (s, 1H), 8.24
6c: 2-Propylthiazolo[4,5-c]quinoline
1
(td, J = 7.1, 3.5 Hz, 4H), 0.89 (dd, J = 9.7, 4.3 Hz, 3H). 13C NMR (d, J = 8.3 Hz, 1H), 7.97 (dd, J = 8.1, 0.9 Hz, 1H), 7.74 (ddd, J =
(126 MHz, CDCl3) δ 172.3, 170.7, 138.0, 132.0, 126.5, 126.3, 8.4, 7.0, 1.4 Hz, 1H), 7.69–7.59 (m, 1H), 3.26–3.11 (m, 2H),
123.5, 123.3, 122.8, 117.6, 37.7, 31.7, 29.1, 25.8, 22.6, 14.2. MS 2.04–1.92 (m, 2H), 1.10 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz,
(ESI) calculated for C16H20N2O2, m/z 272.15, found 273.16 CDCl3) δ 172.8, 147.9, 145.8, 144.3, 140.7, 130.6, 128.8, 127.6,
(M + H)+.
125.0, 123.6, 36.4, 23.3, 13.8. MS (ESI) calculated for
C13H12N2S, m/z 228.07, found 229.08 (M + H)+.
5g: N-(4-Hydroxyquinolin-3-yl)octanamide
6d: 2-Butylthiazolo[4,5-c]quinoline
(263 mg, 73%). 1H NMR (500 MHz, CDCl3) δ 9.25 (d, J = 6.4 Hz,
1
1H), 8.99 (s, 1H), 8.46 (s, 1H), 8.43–8.40 (m, 1H), 7.62 (ddd, J = (108 mg, 73%). H NMR (500 MHz, CDCl3) δ 9.44 (s, 1H), 8.24
8.4, 7.0, 1.4 Hz, 1H), 7.40–7.33 (m, 2H), 2.49–2.44 (m, 2H), (d, J = 8.4 Hz, 1H), 7.96 (dd, J = 8.1, 0.9 Hz, 1H), 7.73 (ddd, J =
1.81–1.73 (m, 2H), 1.41–1.24 (m, 8H), 0.88 (t, J = 6.9 Hz, 3H). 8.4, 5.4, 1.4 Hz, 1H), 7.63 (ddd, J = 8.1, 7.1, 1.1 Hz, 1H),
13C NMR (126 MHz, CDCl3) δ 172.3, 170.8, 138.0, 132.0, 126.6, 3.25–3.19 (m, 2H), 1.97–1.89 (m, 2H), 1.55–1.46 (m, 2H), 1.00
126.3, 123.5, 123.3, 122.8, 117.6, 37.7, 31.9, 29.4, 29.2, 25.9, (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 173.0, 147.9,
1186 | Org. Biomol. Chem., 2013, 11, 1179–1198
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145.8, 144.2, 140.6, 130.6, 128.8, 128.8, 127.6, 125.0, 123.6, 6j: 2-Isobutylthiazolo[4,5-c]quinoline
34.2, 32.0, 22.4, 13.9. MS (ESI) calculated for C14H14N2S, m/z
1
150 mg, 72%. H NMR (500 MHz, MeOD) δ 9.31 (s, 1H), 8.18
(d, J = 8.3 Hz, 1H), 8.11 (dd, J = 8.1, 0.9 Hz, 1H), 7.81 (ddd, J =
8.4, 7.0, 1.4 Hz, 1H), 7.73 (ddd, J = 8.1, 7.1, 1.2 Hz, 1H), 2.30
(m, J = 13.7, 6.8 Hz, 1H), 1.08 (d, J = 6.6 Hz, 6H). 13C NMR
(126 MHz, MeOD) δ 174.5, 148.7, 146.0, 144.7, 142.6, 130.5,
130.4, 129.3, 126.2, 124.7, 43.7, 31.1, 22.6. MS (ESI) calculated
for C14H14N2S, m/z 242.09, found 243.09 (M + H)+.
242.09, found 243.10 (M + H)+.
6e: 2-Pentylthiazolo[4,5-c]quinoline
1
(104 mg, 76%). H NMR (500 MHz, CDCl3) δ 9.45 (s, 1H), 8.24
(d, J = 8.3 Hz, 1H), 7.96 (dd, J = 8.1, 0.9 Hz, 1H), 7.73 (ddd, J =
8.4, 7.0, 1.4 Hz, 1H), 7.63 (ddd, J = 8.1, 7.1, 1.1 Hz, 1H),
3.24–3.17 (m, 2H), 1.95 (dt, J = 15.3, 7.7 Hz, 2H), 1.50–1.36 (m,
4H), 0.93 (t, J = 7.2 Hz, 3H). 13C NMR (126 MHz, CDCl3)
δ 173.1, 147.9, 145.8, 144.2, 140.7, 130.6, 128.8, 127.6, 125.0,
123.6, 34.4, 31.4, 29.6, 22.5, 14.1. MS (ESI) calculated for
C15H16N2S, m/z 256.10, found 257.14 (M + H)+.
6l: 2-Isopentylthiazolo[4,5-c]quinoline
165 mg 75%. 1H NMR (500 MHz, CDCl3) δ 9.44 (s, 1H),
8.26–8.22 (m, 1H), 7.98–7.95 (m, 1H), 7.73 (ddd, J = 8.4, 7.0,
1.4 Hz, 1H), 7.64 (ddd, J = 8.1, 7.0, 1.2 Hz, 1H), 3.26–3.20 (m,
2H), 1.88–1.82 (m, 2H), 1.75 (dd, J = 13.3, 6.7 Hz, 1H), 1.01 (d,
J = 6.6 Hz, 6H). 13C NMR (126 MHz, CDCl3) δ 173.2, 147.9,
145.8, 144.3, 140.6, 130.6, 128.8, 127.6, 125.0, 123.6, 38.8, 32.5,
27.9, 22.5. MS (ESI) calculated for C15H16N2S, m/z 256.10.10,
found 257.10 (M + H)+.
6f: 2-Hexylthiazolo[4,5-c]quinoline
1
(93 mg, 96%). H NMR (500 MHz, CDCl3) δ 9.45 (s, 1H), 8.24
(d, J = 8.4 Hz, 1H), 7.97 (dd, J = 8.1, 0.9 Hz, 1H), 7.74 (ddd, J =
8.4, 7.0, 1.4 Hz, 1H), 7.66–7.62 (m, 1H), 3.25–3.19 (m, 2H),
1.98–1.90 (m, 2H), 1.48 (dd, J = 10.5, 4.5 Hz, 2H), 1.36 (ddd, J =
16.3, 7.0, 4.9 Hz, 4H), 0.90 (dd, J = 9.4, 4.7 Hz, 3H). 13C NMR
(126 MHz, CDCl3) δ 173.1, 147.9, 145.8, 144.3, 140.7, 130.6,
128.8, 127.6, 125.0, 123.6, 34.5, 31.6, 29.9, 28.9, 22.6, 14.2. MS
(ESI) calculated for C16H18N2S, m/z 270.12, found 271.14
(M + H)+.
6o: (S)-2-(2-Methylbutyl)thiazolo[4,5-c]quinoline
170 mg 77%. 1H NMR (500 MHz, CDCl3) δ 9.45 (s, 1H), 8.24 (d,
J = 8.3 Hz, 1H), 7.96 (dd, J = 8.1, 0.9 Hz, 1H), 7.73 (ddd, J = 8.4,
7.0, 1.4 Hz, 1H), 7.63 (ddd, J = 8.1, 7.1, 1.1 Hz, 1H), 3.21 (dd,
J = 14.6, 6.2 Hz, 1H), 3.01 (dd, J = 14.6, 8.2 Hz, 1H), 2.12–2.02
(m, 1H), 1.58–1.48 (m, 1H), 1.40–1.30 (m, 1H), 1.03 (d, J = 6.7
Hz, 3H), 0.98 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3)
δ 172.1, 147.9, 145.8, 144.2, 140.8, 130.6, 128.8, 127.6, 125.0,
123.6, 41.4, 36.3, 29.4, 19.2, 11.5. MS (ESI) calculated for
C15H16N2S, m/z 256.10, found 257.10 (M + H)+.
6g: 2-Heptylthiazolo[4,5-c]quinoline
1
(108 mg, 42%). H NMR (500 MHz, CDCl3) δ 9.44 (s, 1H), 8.23
(d, J = 8.4 Hz, 1H), 7.96 (dd, J = 8.1, 0.9 Hz, 1H), 7.73 (ddd, J =
8.4, 7.0, 1.4 Hz, 1H), 7.65–7.60 (m, 1H), 3.22–3.17 (m, 2H), 1.94
(dt, J = 15.3, 7.6 Hz, 3H), 1.50–1.43 (m, 3H), 1.40–1.35 (m, 2H),
1.31–1.27 (m, 4H), 0.88 (t, J = 7.0 Hz, 3H). 13C NMR (126 MHz,
CDCl3) δ 173.1, 147.9, 145.8, 144.2, 140.6, 130.6, 128.8, 127.6,
125.0, 123.6, 34.5, 31.8, 29.9, 29.2, 29.1, 22.7, 14.2. MS (ESI)
calculated for C17H20N2S, m/z 284.13, found 285.17 (M + H)+.
6p: 2-(3,3,3-Trifluoropropyl)thiazolo[4,5-c]quinoline
180 mg 74%. 1H NMR (500 MHz, CDCl3) δ 9.45 (s, 1H), 8.26 (d,
J = 8.4 Hz, 1H), 7.99–7.95 (m, 1H), 7.76 (ddd, J = 8.4, 7.0, 1.4
Hz, 1H), 7.66 (ddd, J = 8.1, 7.0, 1.2 Hz, 1H), 3.51–3.45 (m, 2H),
2.91–2.80 (m, 2H). 13C NMR (126 MHz, CDCl3) δ 168.0, 147.7,
1
145.8, 144.4, 140.8, 130.7, 129.2, 127.8, 126.4 (q, JCF = 276.5
6h: 2-Nonylthiazolo[4,5-c]quinoline
2
3
Hz), 125.0, 123.4, 33.0 (q, JCF = 29.8 Hz), 27.0 (q, JCF = 3.5
Hz). MS (ESI) calculated for C13H9F3N2S, m/z 282.04, found
283.04 (M + H)+.
1
(157 mg, 52%). H NMR (500 MHz, CDCl3) δ 9.45 (s, 1H), 8.24
(d, J = 8.4 Hz, 1H), 7.96 (d, J = 8.1 Hz, 1H), 7.73 (ddd, J = 8.4,
5.3, 1.4 Hz, 1H), 7.63 (t, J = 7.5 Hz, 1H), 3.23–3.18 (m, 2H), 1.94
(dt, J = 15.4, 7.7 Hz, 2H), 1.51–1.43 (m, 2H), 1.40–1.34 (m, 2H),
1.31–1.24 (m, 8H), 0.87 (t, J = 6.9 Hz, 3H). 13C NMR (126 MHz,
CDCl3) δ 173.1, 147.9, 145.8, 144.2, 130.6, 128.8, 128.7, 127.6,
125.0, 123.6, 120.3, 34.5, 32.0, 29.9, 29.5, 29.4, 29.4, 29.2, 22.8,
14.2. MS (ESI) calculated for C19H24N2S, m/z 312.17, found
313.19 (M + H)+.
6q: 2-(4,4,4-Trifluorobutyl)thiazolo[4,5-c]quinoline
190 mg 75%. 1H NMR (500 MHz, CDCl3) δ 9.46 (s, 1H), 8.26 (d,
J = 8.2 Hz, 1H), 8.00–7.95 (m, 1H), 7.76 (ddd, J = 8.4, 7.0, 1.4
Hz, 1H), 7.66 (ddd, J = 8.1, 7.0, 1.2 Hz, 1H), 3.32 (t, J = 7.2 Hz,
2H), 2.37–2.22 (m, 4H). 13C NMR (126 MHz, CDCl3) δ 170.4,
1
147.9, 145.8, 144.4, 140.7, 130.7, 129.1, 127.8, 127.0 (q, JCF
=
2
276.4 Hz), 125.0, 123.5, 33.9 (q, JCF = 28.9 Hz), 33.0, 21.1 (q,
3JCF = 3.0 Hz). MS (ESI) calculated for C14H11F3N2S, m/z 296.05,
found 297.06 (M + H)+.
6i: 2-Isopropylthiazolo[4,5-c]quinoline
1
164 mg, 83%. H NMR (500 MHz, CDCl3) δ 9.46 (s, 1H), 8.24
(d, J = 8.2 Hz, 1H), 8.01–7.96 (m, 1H), 7.73 (ddd, J = 8.4, 7.0,
1.4 Hz, 1H), 7.64 (ddd, J = 8.1, 7.0, 1.2 Hz, 1H), 3.58–3.48 (m,
1H), 1.56 (d, J = 6.9 Hz, 6H). 13C NMR (126 MHz, CDCl3)
Synthesis of compound 7a: 2-methylthiazolo[4,5-c]quinoline 5-
oxide
δ 179.3, 147.8, 145.9, 144.2, 140.2, 130.6, 128.8, 127.6, 125.0, To a solution of compound 6a (57 mg, 0.28 mmol) in CHCl3
123.7, 34.3, 23.1. MS (ESI) calculated for C13H12N2S, m/z (10 mL), was added m-chloroperoxybenzoic acid (95 mg,
228.10, found 229.08 (M + H)+.
0.43 mmol) and the reaction mixture was stirred at room
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temperature for 18 h. The solvent was then removed under 7g: 2-Heptylthiazolo[4,5-c]quinoline 5-oxide
reduced pressure and the crude residue was purified using
silica gel column chromatography (5% MeOH/CH2Cl2) to
obtain the compound 7a (57 mg, 94%). 1H NMR (500 MHz,
CDCl3) δ 9.13 (s, 1H), 8.89 (d, J = 8.9 Hz, 1H), 7.94 (dd, J = 7.9,
1.1 Hz, 1H), 7.81–7.72 (m, 2H), 2.93 (s, 3H). 13C NMR
(126 MHz, CDCl3) δ 169.7, 147.2, 139.0, 131.5, 131.1, 129.9,
129.7, 125.6, 123.8, 121.6, 20.4. MS (ESI) calculated for
C11H8N2OS, m/z 216.04, found 217.05 (M + H)+.
1
(81 mg, 75%). H NMR (500 MHz, CDCl3) δ 9.13 (s, 1H), 8.88
(dd, J = 8.7, 0.8 Hz, 1H), 7.94 (dd, J = 7.9, 1.1 Hz, 1H),
7.80–7.71 (m, 2H), 3.21–3.15 (m, 2H), 1.91 (dt, J = 15.3, 7.6 Hz,
2H), 1.49–1.42 (m, 2H), 1.41–1.35 (m, 2H), 1.32–1.26 (m, 4H),
0.88 (t, J = 7.0 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 175.3,
147.0, 138.9, 131.6, 130.7, 129.9, 129.6, 125.6, 123.9, 121.6,
34.5, 31.7, 29.8, 29.2, 29.0, 22.7, 14.2. MS (ESI) calculated for
C17H20N2OS, m/z 300.13, found 301.14 (M + H)+.
Compounds 7b–7q were synthesized similarly as compound
7a.
7h: 2-Nonylthiazolo[4,5-c]quinoline 5-oxide
1
(105 mg, 66%). H NMR (500 MHz, CDCl3) δ 9.13 (s, 1H), 8.88
7b: 2-Ethylthiazolo[4,5-c]quinoline 5-oxide
(d, J = 8.5 Hz, 1H), 7.93 (dd, J = 7.9, 1.1 Hz, 1H), 7.81–7.69 (m,
2H), 3.19–3.14 (m, 2H), 1.91 (dt, J = 15.3, 7.6 Hz, 2H), 1.45 (dt,
J = 14.9, 7.0 Hz, 2H), 1.35 (dd, J = 14.2, 6.9 Hz, 2H), 1.29–1.21
(m, 8H), 0.86 (t, J = 6.9 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ
175.3, 147.0, 138.9, 131.6, 130.7, 129.8, 129.6, 125.6, 123.9,
121.5, 34.5, 31.9, 29.8, 29.5, 29.3, 29.2, 22.8, 14.2. MS (ESI) cal-
culated for C19H24N2OS, m/z 328.16, found 329.17 (M + H)+.
1
(81 mg, 88%). H NMR (500 MHz, CDCl3) δ 9.15 (s, 1H), 8.90
(dd, J = 8.6, 0.8 Hz, 1H), 7.95 (dd, J = 8.0, 1.0 Hz, 1H),
7.82–7.72 (m, 2H), 3.23 (q, J = 7.6 Hz, 2H), 1.53 (d, J = 15.1 Hz,
3H). 13C NMR (126 MHz, CDCl3) δ 176.4, 147.1, 139.0, 131.6,
130.7, 129.9, 129.6, 125.6, 123.9, 121.6, 28.0, 13.9. MS (ESI) cal-
culated for C12H10N2OS, m/z 230.05, found 231.06 (M + H)+.
7c: 2-Propylthiazolo[4,5-c]quinoline 5-oxide
7i: 2-Isopropylthiazolo[4,5-c]quinoline 5-oxide
1
(950 mg 88%). H NMR (400 MHz, CDCl3) δ 9.14 (s, 1H), 8.90
1
150 mg, 62%. H NMR (500 MHz, CDCl3) δ 9.15 (s, 1H), 8.90
(d, J = 8.7 Hz, 1H), 7.95 (dd, J = 7.9, 1.1 Hz, 1H), 7.82–7.71 (m,
2H), 3.17 (t, J = 7.6 Hz, 2H), 1.95 (dt, J = 14.9, 7.4 Hz, 2H), 1.09
(t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 175.0, 147.1,
139.0, 131.6, 130.7, 129.9, 129.6, 123.9, 121.6, 36.4, 23.2, 13.8.
MS (ESI) calculated for C13H12N2OS, m/z 244.07, found 245.07
(M + H)+.
(dd, J = 8.7, 1.0 Hz, 1H), 7.99–7.95 (m, 1H), 7.79 (ddd, J = 8.6,
7.0, 1.5 Hz, 1H), 7.77–7.72 (m, 1H), 3.54–3.45 (m, 1H), 1.54 (d,
J = 6.9 Hz, 6H). 13C NMR (126 MHz, CDCl3) δ 181.5, 147.0,
138.9, 131.7, 130.3, 129.9, 129.6, 125.6, 124.0, 121.6, 34.4, 23.0.
MS (ESI) calculated for C13H12N2OS, m/z 244.07.10, found
245.07 (M + H)+.
7d: 2-Butylthiazolo[4,5-c]quinoline 5-oxide
7j: 2-Isobutylthiazolo[4,5-c]quinoline 5-oxide
(82 mg, 75%). 1H NMR (500 MHz, CDCl3) δ 9.13 (s, 1H),
8.90–8.86 (m, 1H), 7.94 (dd, J = 7.9, 1.1 Hz, 1H), 7.80–7.71 (m,
2H), 3.21–3.16 (m, 2H), 1.94–1.86 (m, 2H), 1.53–1.45 (m, 2H),
0.99 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 175.2,
147.1, 139.0, 131.6, 130.6, 129.9, 129.6, 125.6, 123.9, 121.6,
34.2, 31.8, 22.4, 13.9. MS (ESI) calculated for C14H14N2OS, m/z
258.08, found 259.09 (M + H)+.
80 mg, 75%. 1H NMR (500 MHz, MeOD) δ 9.26 (s, 1H), 8.79
(dd, J = 8.6, 0.9 Hz, 1H), 8.25–8.21 (m, 1H), 7.95 (ddd, J = 8.7,
7.1, 1.5 Hz, 1H), 7.90 (ddd, J = 8.2, 7.1, 1.4 Hz, 1H), 3.14 (d, J =
7.2 Hz, 2H), 2.35–2.25 (m, 1H), 1.09 (d, J = 6.6 Hz, 6H). 13C
NMR (126 MHz, MeOD) δ 176.8, 147.8, 139.0, 135.3, 133.5,
131.7, 131.6, 127.2, 125.2, 121.5, 43.7, 31.0, 22.6. MS (ESI) cal-
culated for C14H14N2OS, m/z 258.08.10, found 259.08 (M + H)+.
7e: 2-Pentylthiazolo[4,5-c]quinoline 5-oxide
7k: 2-Neopentylthiazolo[4,5-c]quinoline 5-oxide
1
(90 mg, 87%). H NMR (500 MHz, CDCl3) δ 9.13 (s, 1H), 8.88
80 mg, 75%. 1H NMR (500 MHz, MeOD) δ 9.31 (s, 1H), 8.79
(dd, J = 8.7, 0.9 Hz, 1H), 8.26–8.22 (m, 1H), 7.99–7.88 (m, 2H),
3.16 (s, 2H), 1.13 (s, 9H). 13C NMR (126 MHz, MeOD) δ 174.7,
147.8, 138.8, 135.9, 133.8, 131.8, 131.7, 127.3, 125.1, 121.4,
48.3, 33.2, 29.9. MS (ESI) calculated for C14H16N2OS, m/z
272.10, found 273.10 (M + H)+.
(d, J = 8.1 Hz, 1H), 7.94 (dd, J = 8.0, 1.2 Hz, 1H), 7.81–7.70 (m,
2H), 3.22–3.13 (m, 2H), 1.92 (dt, J = 15.4, 7.7 Hz, 2H),
1.48–1.36 (m, 4H), 0.92 (t, J = 7.1 Hz, 3H). 13C NMR (126 MHz,
CDCl3) δ 175.3, 147.0, 138.9, 131.6, 130.7, 129.9, 129.6, 125.6,
123.9, 121.6, 34.5, 31.3, 29.5, 22.4, 14.0. MS (ESI) calculated for
C15H16N2OS, m/z 272.10, found 273.11 (M + H)+.
7f: 2-Hexylthiazolo[4,5-c]quinoline 5-oxide
7l: 2-Isopentylthiazolo[4,5-c]quinoline 5-oxide
1
1
(65 mg, 76%). H NMR (500 MHz, CDCl3) δ 9.15 (s, 1H), 8.90 110 mg, 69%. H NMR (500 MHz, MeOD) δ 9.23 (s, 1H), 8.77
(d, J = 8.5 Hz, 1H), 7.96 (dd, J = 8.0, 1.0 Hz, 1H), 7.82–7.72 (m, (dd, J = 8.6, 0.9 Hz, 1H), 8.21–8.18 (m, 1H), 7.93 (ddd, J = 8.7,
2H), 3.22–3.17 (m, 2H), 1.92 (dd, J = 15.3, 7.7 Hz, 2H), 7.0, 1.5 Hz, 1H), 7.89 (ddd, J = 8.2, 7.1, 1.3 Hz, 1H), 3.27 (dd,
1.51–1.44 (m, 2H), 1.40–1.30 (m, 4H), 0.90 (t, J = 7.1 Hz, 3H). J = 8.4, 7.4 Hz, 2H), 1.88–1.82 (m, 2H), 1.78–1.69 (m, 1H), 1.02
13C NMR (126 MHz, CDCl3) δ 175.3, 147.1, 139.0, 131.7, 130.8, (d, J = 6.6 Hz, 6H). 13C NMR (126 MHz, MeOD) δ 178.1, 147.7,
129.9, 129.6, 125.6, 123.9, 121.6, 34.5, 31.6, 29.8, 28.9, 22.6, 138.9, 135.1, 133.5, 131.6, 131.6, 127.1, 125.1, 121.5, 39.6, 33.1,
14.2. MS (ESI) calculated for C16H18N2OS, m/z 286.11, found 28.8, 22.6. MS (ESI) calculated for C15H16N2OS, m/z 272.10,
287.12 (M + H)+.
found 273.10 (M + H)+.
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7m: 2-(3,3-Dimethylbutyl)thiazolo[4,5-c]quinoline 5-oxide
solvent was then removed under vacuum and the residue was
dissolved in anhydrous methanol (5 mL). Excess of sodium
methoxide was added and the reaction mixture was refluxed
for 2 h. After evaporating solvents under reduced pressure, the
crude residue was purified using silica gel column chromato-
graphy (7% MeOH/CH2Cl2) to obtain compound 8a as a white
80 mg, 76%. 1H NMR (500 MHz, MeOD) δ 9.22 (s, 1H), 8.77
(dd, J = 8.7, 0.9 Hz, 1H), 8.21–8.18 (m, 1H), 7.93 (ddd, J = 8.7,
7.1, 1.5 Hz, 1H), 7.89 (ddd, J = 8.2, 7.1, 1.3 Hz, 1H), 3.27–3.22
(m, 2H), 1.89–1.84 (m, 2H), 1.04 (s, 9H). 13C NMR (126 MHz,
MeOD) δ 178.7, 147.7, 138.9, 135.1, 133.4, 131.6, 131.6, 127.1,
125.1, 121.4, 44.7, 31.5, 30.9, 29.5. MS (ESI) calculated for
C16H18N2OS, m/z 286.11, found 287.11 (M + H)+.
1
solid (41 mg, 82%). H NMR (400 MHz, CDCl3) δ 7.77 (dd, J =
8.4, 0.5 Hz, 1H), 7.72 (dd, J = 8.0, 1.0 Hz, 1H), 7.55 (ddd, J =
8.4, 7.1, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.2 Hz, 1H), 5.61 (s,
2H), 2.89 (s, 3H). 13C NMR (126 MHz, CDCl3) δ 165.9, 151.5,
144.8, 141.0, 138.1, 129.1, 126.8, 124.8, 123.2, 120.1, 20.1. MS
(ESI) calculated for C11H9N3S, m/z 215.05, found 216.06 (M +
H)+.
7n: (S)-2-(sec-Butyl)thiazolo[4,5-c]quinoline 5-oxide
1
117 mg, 78%. H NMR (500 MHz, MeOD) δ 9.23 (s, 1H), 8.77
(dd, J = 8.6, 1.0 Hz, 1H), 8.22–8.18 (m, 1H), 7.93 (ddd, J = 8.7,
7.1, 1.5 Hz, 1H), 7.88 (ddd, J = 8.2, 7.1, 1.4 Hz, 1H), 3.35 (dd, J
= 13.9, 6.9 Hz, 1H), 2.04–1.94 (m, 1H), 1.92–1.82 (m, 1H), 1.52
(d, J = 6.9 Hz, 3H), 1.02 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz,
MeOD) δ 183.1, 147.7, 138.9, 134.7, 133.5, 131.6, 131.6, 127.1,
125.2, 121.5, 42.3, 31.6, 20.9, 12.0. MS (ESI) calculated for
C14H14N2OS, m/z 258.09, found 259.08 (M + H)+.
Compounds 8b–8q were synthesized similarly as compound
8a.
8b: 2-Ethylthiazolo[4,5-c]quinolin-4-amine
White solid (36 mg, 47%). 1H NMR (400 MHz, CDCl3) δ 7.77
(dd, J = 8.4, 0.5 Hz, 1H), 7.74 (dd, J = 8.0, 1.1 Hz, 1H), 7.56
(ddd, J = 8.4, 7.1, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.1 Hz,
1H), 5.58 (s, 2H), 3.20 (q, J = 7.6 Hz, 2H), 1.51 (t, J = 7.6 Hz,
3H). 13C NMR (126 MHz, CDCl3) δ 172.6, 151.6, 144.8, 140.4,
138.0, 129.0, 126.8, 124.8, 123.2, 120.2, 27.8, 14.1. MS (ESI) cal-
culated for C12H11N3S, m/z 229.07, found 230.08 (M + H)+.
7o: (S)-2-(2-Methylbutyl)thiazolo[4,5-c]quinoline 5-oxide
1
103 mg, 74%. H NMR (500 MHz, MeOD) δ 9.24 (s, 1H), 8.78
(dd, J = 8.7, 0.8 Hz, 1H), 8.23–8.19 (m, 1H), 7.94 (ddd, J = 8.7,
7.1, 1.5 Hz, 1H), 7.89 (dd, 1H), 3.26 (dd, J = 14.7, 6.2 Hz, 1H),
3.06 (dd, J = 14.7, 8.0 Hz, 1H), 2.14–2.03 (m, 1H), 1.60–1.50 (m,
1H), 1.44–1.30 (m, 1H), 1.04 (d, J = 6.7 Hz, 3H), 1.00 (t, J = 7.4
Hz, 3H). 13C NMR (126 MHz, MeOD) δ 177.0, 147.8, 139.0,
135.3, 133.5, 131.7, 131.6, 127.1, 125.1, 121.5, 41.8, 37.4, 30.2,
19.4, 11.6. MS (ESI) calculated for C15H16N2OS, m/z 272.10,
found 273.10 (M + H)+.
8c: 2-Propylthiazolo[4,5-c]quinolin-4-amine
1
White solid (450 mg, 90%). H NMR (400 MHz, CDCl3) δ 7.77
(d, J = 8.4 Hz, 1H), 7.73 (dd, J = 8.0, 1.3 Hz, 1H), 7.55 (ddd, J =
8.4, 7.1, 1.4 Hz, 1H), 7.34–7.27 (m, 1H), 5.67 (s, 2H), 3.13 (t, J =
7.6 Hz, 2H), 1.94 (dd, J = 15.0, 7.4 Hz, 2H), 1.09 (t, J = 7.4 Hz,
3H). 13C NMR (126 MHz, CDCl3) δ 171.2, 151.7, 144.7, 140.5,
138.0, 129.0, 126.8, 124.8, 123.1, 120.2, 36.2, 23.4, 13.8. MS
(ESI) calculated for C13H13N3S, m/z 243.08, found 244.09
(M + H)+.
7p: 2-(3,3,3-Trifluoropropyl)thiazolo[4,5-c]quinoline 5-oxide
1
120 mg, 75%. H NMR (500 MHz, MeOD) δ 9.24 (s, 1H), 8.76
(dd, J = 8.8, 0.7 Hz, 1H), 8.17 (dd, 1H), 7.93 (ddd, J = 8.6, 7.0,
1.4 Hz, 1H), 7.88 (ddd, J = 8.2, 7.1, 1.3 Hz, 1H), 3.54 (t, 2H),
2.99–2.87 (m, 2H). 13C NMR (126 MHz, MeOD) δ 173.8, 147.6,
1
139.1, 135.2, 133.6, 131.8, 131.6, 128.1 (q, JCF = 275.6 Hz),
8d: 2-Butylthiazolo[4,5-c]quinolin-4-amine
2
3
127.1, 125.0, 121.5, 33.2 (q, JCF = 29.7 Hz), 27.7 (q, JCF = 3.5
Hz). MS (ESI) calculated for C13H9F3N2OS, m/z 298.04, found
299.04 (M + H)+.
White solid (25 mg, 32%). 1H NMR (400 MHz, CDCl3) δ 7.77
(dd, J = 8.4, 0.6 Hz, 1H), 7.73 (dd, J = 8.0, 1.0 Hz, 1H), 7.56
(ddd, J = 8.5, 7.1, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.2 Hz,
1H), 5.56 (s, 2H), 3.20–3.13 (m, 2H), 1.90 (dt, J = 15.3, 7.6 Hz,
2H), 1.50 (dq, J = 14.7, 7.4 Hz, 2H), 1.00 (t, J = 7.4 Hz, 3H). 13C
NMR (126 MHz, CDCl3) δ 171.5, 151.6, 144.7, 140.5, 138.0,
129.0, 126.8, 124.8, 123.2, 120.2, 34.0, 32.0, 22.4, 13.9. MS (ESI)
calculated for C14H15N3S, m/z 257.10, found 258.11 (M + H)+.
7q: 2-(4,4,4-Trifluorobutyl)thiazolo[4,5-c]quinoline 5-oxide
128 mg, 81%. 1H NMR (500 MHz, MeOD) δ 9.26 (s, 1H),
8.80–8.74 (m, 1H), 8.22–8.18 (m, 1H), 7.94 (ddd, J = 8.6, 7.1,
1.5 Hz, 1H), 7.89 (ddd, J = 8.2, 7.1, 1.3 Hz, 1H), 3.36 (t, J = 7.5
Hz, 2H), 2.48–2.34 (m, 2H), 2.27–2.17 (m, 2H). 13C NMR
(126 MHz, MeOD) δ 176.0, 147.8, 139.0, 135.2, 133.6, 131.7,
1
8e: 2-Pentylthiazolo[4,5-c]quinolin-4-amine
131.6, 128.6 (q, JCF = 275.4 Hz), 127.1, 125.1, 121.5, 33.6 (q,
2JCF = 29.0 Hz), 33.5, 22.6 (q, 3JCF = 3.2 Hz). MS (ESI) calculated
for C14H11F3N2OS, m/z 312.05, found 313.05 (M + H)+.
White solid (38 mg, 45%). 1H NMR (400 MHz, CDCl3) δ 7.77
(d, J = 8.1 Hz, 1H), 7.73 (dd, J = 8.0, 1.1 Hz, 1H), 7.55 (ddd, J =
8.4, 7.1, 1.4 Hz, 1H), 7.34–7.29 (m, 1H), 5.59 (s, 2H), 3.18–3.12
(m, 2H), 1.96–1.86 (m, 2H), 1.50–1.35 (m, 4H), 0.94 (t, J = 7.1
Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 171.5, 151.6, 144.7,
Synthesis of compound 8a: 2-methylthiazolo[4,5-c]quinolin-4-
amine
Compound 7a (50 mg, 0.23 mmol) was dissolved in anhydrous 140.5, 138.0, 129.0, 126.8, 124.8, 123.2, 120.2, 34.3, 31.4, 29.7,
CH2Cl2 (5 mL). Benzoyl isocyanate (68 mg, 0.46 mmol) was 22.5, 14.1 MS (ESI) calculated for C15H17N3S, m/z 271.11,
added to the reaction mixture and refluxed for 30 min. The found 272.12 (M + H)+.
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8f: 2-Hexylthiazolo[4,5-c]quinolin-4-amine
8k: 2-Neopentylthiazolo[4,5-c]quinolin-4-amine
White solid (25 mg, 44%). 1H NMR (400 MHz, CDCl3) δ 7.77 White solid (45 mg, 76%). 1H NMR (500 MHz, CDCl3) δ 7.77
(dd, J = 8.4, 0.5 Hz, 1H), 7.73 (dd, J = 8.0, 1.0 Hz, 1H), 7.55 (dd, J = 8.4, 0.5 Hz, 1H), 7.74 (dd, J = 8.0, 1.0 Hz, 1H), 7.55
(ddd, J = 8.4, 7.1, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.1 Hz, (ddd, J = 8.4, 5.5, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.1 Hz,
1H), 5.56 (s, 2H), 3.21–3.11 (m, 2H), 1.91 (dt, J = 15.3, 7.5 Hz, 1H), 5.60 (s, 2H), 3.04 (s, 2H), 1.10 (s, 9H). 13C NMR (126 MHz,
2H), 1.46 (dt, J = 9.2, 7.0 Hz, 2H), 1.39–1.30 (m, 4H), 0.91 (t, J = CDCl3) δ 168.1, 151.7, 144.8, 140.7, 138.2, 129.1, 126.8, 124.9,
7.1 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 171.6, 151.6, 144.7, 123.2, 120.1, 47.8, 32.3, 29.7. MS (ESI) calculated for
140.51, 138.0, 129.1, 126.8, 124.8, 123.2, 120.2, 34.3, 31.6, 30.0, C14H17N3S, m/z 271.11, found 272.11 (M + H)+.
29.0, 22.6, 14.2. MS (ESI) calculated for C16H19N3S, m/z 285.13,
found 286.14 (M + H)+.
8l: 2-Isopentylthiazolo[4,5-c]quinolin-4-amine
White solid (40 mg, 68%). 1H NMR (500 MHz, CDCl3) δ 7.77
(dd, J = 8.4, 0.5 Hz, 1H), 7.73 (dd, J = 8.0, 1.0 Hz, 1H), 7.55
(ddd, J = 8.4, 7.1, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.1 Hz,
1H), 5.61 (s, 2H), 3.20–3.13 (m, 2H), 1.84–1.78 (m, 2H),
1.77–1.69 (m, 1H), 1.00 (d, J = 6.5 Hz, 6H). 13C NMR (126 MHz,
CDCl3) δ 171.7, 151.6, 144.7, 140.5, 138.0, 129.0, 126.8, 124.8,
123.2, 120.2, 38.9, 32.3, 27.9, 22.5. MS (ESI) calculated for
C15H16N2OS, m/z 271.11, found 272.11 (M + H)+.
8g: 2-Heptylthiazolo[4,5-c]quinolin-4-amine
White solid (34 mg, 42%). 1H NMR (400 MHz, CDCl3) δ 7.77
(dd, J = 8.4, 0.5 Hz, 1H), 7.73 (dd, J = 8.0, 1.0 Hz, 1H), 7.55
(ddd, J = 8.4, 7.1, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.2 Hz,
1H), 5.56 (s, 2H), 3.20–3.08 (m, 2H), 1.91 (dt, J = 15.3, 7.6 Hz,
2H), 1.50–1.42 (m, 2H), 1.38 (ddd, J = 14.0, 8.2, 5.2 Hz, 2H),
1.30 (dt, J = 11.5, 3.9 Hz, 4H), 0.89 (t, J = 6.9 Hz, 3H). 13C NMR
(126 MHz, CDCl3) δ 171.6, 151.6, 144.7, 140.5, 138.0, 129.0,
126.8, 124.8, 123.2, 120.2, 34.3, 31.8, 30.0, 29.2, 29.1, 22.8,
14.2. MS (ESI) calculated for C17H21N3S, m/z 299.15, found
300.16 (M + H)+.
8m: 2-(3,3-Dimethylbutyl)thiazolo[4,5-c]quinolin-4-amine
White solid (45 mg, 65%). 1H NMR (500 MHz, CDCl3) δ 7.77
(dd, J = 8.4, 0.5 Hz, 1H), 7.74–7.70 (m, 1H), 7.55 (ddd, J = 8.4,
7.0, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.1 Hz, 1H), 5.64 (s,
2H), 3.17–3.09 (m, 2H), 1.86–1.79 (m, 2H), 1.02 (s, 9H). 13C
NMR (126 MHz, CDCl3) δ 172.2, 151.6, 144.7, 140.4, 138.0,
129.0, 126.8, 124.8, 123.1, 121.0, 44.1, 30.8, 30.1, 29.3. MS (ESI)
calculated for C16H19N3S, m/z 285.13, found 286.14 (M + H)+.
8h: 2-Nonylthiazolo[4,5-c]quinolin-4-amine
1
(26 mg, 26%). H NMR (400 MHz, CDCl3) δ 7.77 (dd, J = 8.4,
0.5 Hz, 1H), 7.73 (dd, J = 8.0, 1.0 Hz, 1H), 7.55 (ddd, J = 8.5,
7.1, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.2 Hz, 1H), 5.57 (s,
2H), 3.21–3.09 (m, 2H), 1.91 (dt, J = 15.3, 7.6 Hz, 2H), 1.46
(ddd, J = 14.9, 8.7, 6.5 Hz, 2H), 1.36 (dd, J = 13.7, 6.7 Hz, 2H),
1.28 (s, 8H), 0.88 (t, J = 6.9 Hz, 3H). 13C NMR (126 MHz,
CDCl3) δ 171.6, 151.6, 144.7, 140.5, 138.0, 129.0, 126.8, 124.8,
123.2, 120.2, 34.3, 32.0, 30.0, 29.6, 29.4, 29.4, 29.2, 22.8, 14.3.
MS (ESI) calculated for C19H25N3S, m/z 327.18, found 328.19
(M + H)+.
8n: (S)-2-(sec-Butyl)thiazolo[4,5-c]quinolin-4-amine
White solid (45 mg, 76%). 1H NMR (500 MHz, CDCl3) δ
7.79–7.75 (m, 1H), 7.75–7.72 (m, 1H), 7.55 (ddd, J = 8.4, 7.0,
1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.2 Hz, 1H), 5.63 (s, 2H),
3.30–3.21 (m, 1H), 2.00–1.90 (m, 1H), 1.88–1.76 (m, 1H), 1.49
(d, J = 6.9 Hz, 3H), 1.00 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz,
CDCl3) δ 177.0, 151.7, 144.7, 140.0, 137.9, 129.0, 126.8, 124.8,
123.1, 120.3, 41.0, 30.9, 21.0, 11.9. MS (ESI) calculated for
C14H15N3S, m/z 257.10, found 258.10 (M + H)+.
8i: 2-Isopropylthiazolo[4,5-c]quinolin-4-amine
White solid (40 mg, 67%). 1H NMR (500 MHz, CDCl3) δ 7.77
(dd, J = 8.4, 0.5 Hz, 1H), 7.75–7.71 (m, 1H), 7.55 (ddd, J = 8.4,
7.0, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.2 Hz, 1H), 5.66 (s,
2H), 3.51–3.41 (m, 1H), 1.52 (d, J = 6.9 Hz, 6H). 13C NMR
(126 MHz, CDCl3) δ 177.7, 151.8, 144.7, 140.0, 137.9, 129.0,
126.8, 124.8, 123.1, 120.3, 34.2, 23.2. MS (ESI) calculated for
C13H13N3S, m/z 243.09.10, found 244.09 (M + H)+.
8o: (S)-2-(2-Methylbutyl)thiazolo[4,5-c]quinolin-4-amine
White solid (46 mg, 77%). 1H NMR (500 MHz, CDCl3) δ 7.77
(dd, J = 8.4, 0.5 Hz, 1H), 7.73 (dd, J = 8.0, 1.0 Hz, 1H), 7.55
(ddd, J = 8.4, 7.1, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.1 Hz,
1H), 5.66 (s, 2H), 3.14 (dd, J = 14.6, 6.2 Hz, 1H), 2.96 (dd, J =
14.6, 8.1 Hz, 1H), 2.08–1.97 (m, 1H), 1.57–1.47 (m, 1H),
1.38–1.28 (m, 1H), 1.02 (d, J = 6.7 Hz, 3H), 0.97 (t, J = 7.4 Hz,
3H). 13C NMR (126 MHz, CDCl3) δ 170.5, 151.7, 144.7, 140.6,
138.1, 129.0, 126.8, 124.8, 123.1, 120.2, 41.2, 36.3, 29.3, 19.2,
11.5. MS (ESI) calculated for C15H17N3S, m/z 271.11, found
272.11 (M + H)+.
8j: 2-Isobutylthiazolo[4,5-c]quinolin-4-amine
White solid (38 mg, 77%). 1H NMR (500 MHz, CDCl3) δ 7.77
(dd, J = 8.4, 0.5 Hz, 1H), 7.73 (dd, J = 8.0, 1.0 Hz, 1H), 7.55
(ddd, J = 8.4, 7.1, 1.5 Hz, 1H), 7.31 (ddd, J = 8.1, 7.1, 1.1 Hz,
1H), 5.63 (s, 2H), 3.02 (d, J = 7.2 Hz, 2H), 2.30–2.20 (m, 1H),
8p: 2-(3,3,3-Trifluoropropyl)thiazolo[4,5-c]quinolin-4-amine
1.06 (d, J = 6.6 Hz, 6H). 13C NMR (126 MHz, CDCl3) δ 170.3, White solid (70 mg, 78%). 1H NMR (500 MHz, CDCl3) δ
151.7, 144.8, 140.6, 138.1, 129.1, 126.8, 124.9, 123.1, 120.2, 7.80–7.76 (m, 1H), 7.73 (dd, J = 8.0, 1.0 Hz, 1H), 7.58 (ddd, J =
43.1, 30.0, 22.5. MS (ESI) calculated for C14H15N3S, m/z 257.10, 8.4, 7.1, 1.4 Hz, 1H), 7.35–7.30 (m, 1H), 5.63 (s, 2H), 3.45–3.39
found 258.10 (M + H)+.
(m, 2H), 2.87–2.76 (m, 2H). 13C NMR (126 MHz, CDCl3) δ
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166.5, 151.6, 145.0, 140.8, 137.9, 129.4, 126.9, 126.4 (q, JCF
Paper
1
=
sodium nitrite (20 mg, 0.29 mmol) and the reaction mixture
2
276.5 Hz), 124.9, 123.4, 119.9, 33.1 (q, JCF = 29.8 Hz), 26.8 (q, was stirred at room temperature for 1 h. Sodium azide
3JCF = 3.4 Hz). MS (ESI) calculated for C13H10F3N3S, m/z 297.05, (18.8 mg, 0.29 mmol) was then added to the reaction mixture
found 298.05 (M + H)+.
and stirred for additional 1 h. The reaction mixture was neu-
tralized with 1 N NaOH and extracted with EtOAc (3 × 10 mL).
The combined organic layer was dried over Na2SO4, concen-
8q: 2-(4,4,4-Trifluorobutyl)thiazolo[4,5-c]quinolin-4-amine
White solid (50 mg, 72%). 1H NMR (500 MHz, CDCl3) δ 7.77 trated under reduced pressure, and the crude compound was
(dd, J = 8.4, 0.4 Hz, 1H), 7.73 (dd, J = 8.0, 1.1 Hz, 1H), 7.57 purified on silica gel column chromatography (5% MeOH/
(ddd, J = 8.4, 7.1, 1.4 Hz, 1H), 7.32 (ddd, J = 8.1, 7.1, 1.1 Hz, CH2Cl2) to obtain compound 11 as a brown solid (18 mg,
1H), 5.62 (s, 2H), 3.25 (t, J = 7.2 Hz, 2H), 2.36–2.26 (m, 2H), 67%). 1H NMR (400 MHz, CDCl3) δ 7.76 (d, J = 8.9 Hz, 1H),
2.26–2.18 (m, 2H). 13C NMR (126 MHz, CDCl3) δ 168.8, 151.6, 7.31 (d, J = 2.5 Hz, 1H), 7.26–7.23 (m, 1H), 5.60 (s, 2H),
1
144.9, 140.6, 138.1, 129.3, 127.0 (q, JCF = 276.4 Hz), 126.9, 3.17–3.12 (m, 2H), 2.01–1.90 (m, 2H), 1.09 (t, J = 7.4 Hz, 3H).
3
124.9, 123.3, 120.0, 33.1 (q, 2JCF = 29.7 Hz), 32.7, 21.9 (q, JCF
=
13C NMR (126 MHz, CDCl3) δ 172.0, 151.3, 139.5, 138.5, 134.8,
3.0 Hz). MS (ESI) calculated for C14H12F3N3S, m/z 311.07, 128.4, 121.0, 120.8, 113.5, 36.2, 23.4, 13.8. MS (ESI) calculated
found 312.07 (M + H)+.
for C13H12N6S, m/z 284.08, found 285.09 (M + H)+.
Synthesis of compound 9: 8-nitro-2-propylthiazolo[4,5-c]-
Synthesis of compound 12a: 2-(1-(4-amino-2-propylthiazolo-
quinolin-4-amine
[4,5-c]quinolin-8-yl)-1H-1,2,3-triazol-4-yl)propan-2-ol
To a stirred solution of compound 8c (200 mg, 0.82 mmol) in To a stirred solution of compound 11 (12 mg, 0.042 mmol) in
H2SO4 (0.5 mL), was added HNO3 (0.5 mL) and the reaction THF (1 mL), were added CuSO4·5H2O (1.3 mg, 0.005 mmol, in
mixture was stirred at room temperature for 1 h. The reaction 0.5 mL water), sodium ascorbate (2.1 mg, 0.010 mmol, in
mixture was neutralized with 1 N NaOH and extracted with 0.5 mL water), 2-methylbut-3-yn-2-ol (8.4 μL, 0.084 mmol), and
EtOAc (3 × 15 mL). The combined organic layer was dried over the reaction mixture stirred at room temperature for 1 h. The
Na2SO4, concentrated under reduced pressure, and the crude reaction mixture was diluted with water and extracted with
residue was purified using silica gel column chromatography EtOAc (3 × 10 mL). The combined organic layer was dried over
(5% MeOH/CH2Cl2) to obtain compound 9 as yellow solid Na2SO4, concentrated under reduced pressure, and the crude
(171 mg, 72%). 1H NMR (400 MHz, CDCl3) δ 8.68 (d, J = 2.5 Hz, residue was purified using silica gel column chromatography
1H), 8.35 (dd, J = 9.2, 2.6 Hz, 1H), 7.80 (d, J = 9.2 Hz, 1H), 6.09 (10% MeOH/CH2Cl2) to obtain compound 11 as a white solid
1
(s, 2H), 3.17 (dd, J = 8.7, 6.4 Hz, 2H), 1.97 (dd, J = 15.0, 7.5 Hz, (14 mg, 92%). H NMR (400 MHz, CDCl3) δ 8.12 (d, J = 2.0 Hz,
2H), 1.11 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 1H), 7.97 (s, 1H), 7.91–7.81 (m, 2H), 5.74 (s, 2H), 3.49 (s, 1H),
173.3, 153.6, 148.3, 142.5, 141.2, 138.7, 127.3, 123.2, 121.5, 3.19–3.13 (m, 2H), 1.96 (dd, J = 15.0, 7.5 Hz, 2H), 1.74 (s, 6H),
119.1, 36.2, 23.3, 13.8. MS (ESI) calculated for C13H12N4O2S, 1.10 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 172.5,
m/z 288.07, found 289.08 (M + H)+.
156.5, 152.2, 144.5, 140.2, 138.7, 132.0, 128.3, 121.5, 120.3,
117.8, 116.4, 68.9, 36.2, 30.7, 23.4, 13.8. MS (ESI) calculated for
C18H20N6OS, m/z 368.14, found 369.15 (M + H)+.
Synthesis of compound 10: 2-propylthiazolo[4,5-c]quinoline-
4,8-diamine
Compounds 12b–12f were synthesized similarly as com-
To a stirred solution of compound 9 (200 mg, 0.69 mmol) in pound 12a.
methanol (5 mL), were added zinc dust (453 mg, 6.94 mmol)
and ammonium formate (437 mg, 6.94 mmol), and the reac-
tion mixture stirred at room temperature for 30 min. The reac-
12b: (1-(4-Amino-2-propylthiazolo[4,5-c]quinolin-8-yl)-1H-1,2,3-
triazol-4-yl)methanol
1
tion mixture was then filtered through celite, the filtrate White solid (15 mg, 83%). H NMR (500 MHz, DMSO) δ 8.84
concentrated under reduced pressure, and purified over silica (s, 1H), 8.33 (d, J = 2.5 Hz, 1H), 8.06 (dd, J = 9.0, 2.5 Hz, 1H),
gel column chromatography (10% MeOH/CH2Cl2) to afford 7.76 (d, J = 9.0 Hz, 1H), 7.13 (s, 2H), 5.37 (t, J = 5.6 Hz, 1H),
compound 10 as a yellow solid (185 mg, 88%). 1H NMR 4.64 (d, J = 5.9 Hz, 2H), 3.19 (t, J = 7.5 Hz, 2H), 1.88 (dt, J =
(400 MHz, CDCl3) δ 8.71 (s, 1H), 8.46 (s, 2H), 7.70 (d, J = 8.9 14.8, 7.4 Hz, 2H), 1.04 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz,
Hz, 1H), 7.00 (dd, J = 8.9, 2.5 Hz, 1H), 6.87 (d, J = 2.4 Hz, 1H), DMSO) δ 171.8, 152.7, 149.2, 144.5, 139.1, 138.2, 130.8, 127.3,
3.17–3.09 (m, 2H), 1.99–1.88 (m, 2H), 1.08 (t, J = 7.4 Hz, 3H). 121.2, 120.9, 118.9, 115.2, 55.1, 35.2, 22.8, 13.5. MS (ESI) calcu-
13C NMR (126 MHz, CDCl3) δ 173.1, 170.3, 149.2, 143.9, 142.1, lated for C16H16N6OS, m/z 340.11, found 341.11 (M + H)+.
137.1, 129.5, 121.9, 120.7, 118.7, 107.4, 36.1, 23.2, 13.8. MS
12c: 8-(4-Phenyl-1H-1,2,3-triazol-1-yl)-2-propylthiazolo[4,5-c]-
(ESI) calculated for C13H14N4S, m/z 258.09, found 259.10
quinolin-4-amine
(M + H)+.
1
Solid (19 mg, 93%). H NMR (500 MHz, DMSO) δ 9.44 (s, 1H),
Synthesis of compound 11: 8-azido-2-propylthiazolo[4,5-c]-
8.36 (d, J = 2.5 Hz, 1H), 8.12 (dd, J = 9.0, 2.5 Hz, 1H), 7.97 (dd,
quinolin-4-amine
J = 8.2, 1.1 Hz, 2H), 7.80 (d, J = 9.0 Hz, 1H), 7.53 (dd, J = 10.6,
To a stirred solution of compound 10 (25 mg, 0.096 mmol) in 4.8 Hz, 2H), 7.44–7.37 (m, 1H), 7.17 (s, 2H), 3.21 (t, J = 7.5 Hz,
a 1 : 1 mixture of acetic acid and water (2 mL), was added 2H), 1.90 (dd, J = 14.9, 7.4 Hz, 2H), 1.04 (t, J = 7.4 Hz, 3H). 13C
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NMR (126 MHz, DMSO) δ 171.8, 152.8, 147.3, 144.7, 139.0, 134.4, 130.7, 127.4, 123.4, 121.2, 118.9, 115.5, 35.23 22.8, 13.5.
138.2, 130.6, 130.4, 129.1, 128.3, 127.4, 125.3, 120.9, 119.8, MS (ESI) calculated for C15H14N6S, m/z 310.10, found 311.10
118.9, 115.3, 35.2, 22.8, 13.5. MS (ESI) calculated for (M + H)+.
C21H18N6S, m/z 386.13, found 387.14 (M + H)+.
Synthesis of compound 13a
12d: 2-Propyl-8-(4-propyl-1H-1,2,3-triazol-1-yl)thiazolo[4,5-c]-
quinolin-4-amine
N8-Hexyl-2-propylthiazolo[4,5-c]quinoline-4,8-diamine. To
stirred solution of compound 10 (25 mg, 0.096 mmol) in DMF
a
1
Yellow solid (16 mg, 86%). H NMR (500 MHz, DMSO) δ 8.72 (2 mL) were added 1-iodohexane (20.5 mg, 0.096 mmol) and
(s, 1H), 8.28 (d, J = 2.4 Hz, 1H), 8.04 (dd, J = 9.0, 2.5 Hz, 1H), potassium carbonate (26.7 mg, 0.19 mmol), and the reaction
7.75 (d, J = 9.0 Hz, 1H), 7.12 (s, 2H), 3.19 (t, J = 7.5 Hz, 2H), mixture stirred at room temperature for 4 h. The reaction
2.71 (t, J = 7.5 Hz, 2H), 1.89 (dd, J = 14.9, 7.4 Hz, 2H), 1.72 (dd, mixture was diluted with water and extracted with EtOAc (3 ×
J = 14.9, 7.4 Hz, 2H), 1.04 (t, J = 7.4 Hz, 3H), 0.98 (t, J = 7.4 Hz, 10 mL). The organic layer was dried over Na2SO4, concentrated
3H). 13C NMR (126 MHz, DMSO) δ 171.8, 152.7, 148.0, 144.4, under reduced pressure, and the crude compound was puri-
139.0, 138.2, 130.9, 127.3, 120.8, 120.3, 118.9, 115.0, 35.2, 27.1, fied using silica gel column chromatography (5% MeOH/
22.8, 22.1, 13.7, 13.5. MS (ESI) calculated for C18H20N6S, m/z CH2Cl2) to obtain 13a as a solid (16 mg, 61%). 1H NMR
352.15, found 353.15 (M + H)+.
(500 MHz, CDCl3) δ 7.69 (d, J = 8.9 Hz, 1H), 6.96 (dd, J = 8.9,
2.6 Hz, 1H), 6.67 (d, J = 2.5 Hz, 1H), 6.15 (s, 2H), 3.87 (s, 1H),
3.18 (t, J = 7.1 Hz, 2H), 3.15–3.11 (m, 2H), 1.99–1.89 (m, 2H),
1.68 (dt, J = 14.7, 7.2 Hz, 2H), 1.49–1.41 (m, 2H), 1.35 (td, J =
12e: 8-(4-Pentyl-1H-1,2,3-triazol-1-yl)-2-propylthiazolo[4,5-c]-
quinolin-4-amine
1
Solid (19 mg, 91%). H NMR (500 MHz, DMSO) δ 8.71 (s, 1H), 7.2, 3.7 Hz, 4H), 1.09 (t, J = 7.4 Hz, 3H), 0.92 (t, J = 7.1 Hz, 3H).
8.27 (d, J = 2.5 Hz, 1H), 8.04 (dd, J = 9.0, 2.5 Hz, 1H), 7.75 (d, 13C NMR (126 MHz, CDCl3) δ 172.1, 148.2, 145.5, 141.2, 137.4,
J = 9.0 Hz, 1H), 7.12 (s, 2H), 3.19 (t, J = 7.5 Hz, 2H), 2.72 (t, J = 124.8, 120.3, 119.8, 102.9, 44.3, 36.2, 31.8, 29.4, 27.0, 23.3,
7.6 Hz, 2H), 1.89 (dd, J = 14.9, 7.4 Hz, 2H), 1.69 (dt, J = 15.3, 22.8, 14.2, 13.8. MS (ESI) calculated for C19H26N4S, m/z 342.19,
7.6 Hz, 2H), 1.42–1.26 (m, 6H), 1.04 (t, J = 7.4 Hz, 3H), 0.87 found 343.20 (M + H)+.
(dd, J = 9.2, 4.9 Hz, 3H). 13C NMR (126 MHz, DMSO) δ 171.8,
152.7, 148.2, 144.4, 139.0, 138.2, 130.9, 127.3, 120.8, 120.2,
Synthesis of compound 13b: N-(4-amino-2-propylthiazolo[4,5-c]-
quinolin-8-yl)butyramide
118.9, 114.9, 35.2, 31.1, 28.8, 28.3, 25.1, 22.8, 22.1, 14.0, 13.5.
MS (ESI) calculated for C21H26N6S, m/z 394.19, found 395.20 To a stirred solution of compound 10 (20 mg, 0.077 mmol) in
(M + H)+.
CH2Cl2 (2 mL) were added triethylamine (30 μL, 0.164 mmol)
and butyryl chloride (8.7 μL, 0.082); the reaction mixture was
stirred at room temperature for 1 h. The reaction mixture was
then diluted with water and extracted with EtOAc (3 × 10 mL).
12f: 2-Propyl-8-(4-(trimethylsilyl)-1H-1,2,3-triazol-1-yl)thiazolo-
[4,5-c]quinolin-4-amine
1
White solid (18 mg, 90%). H NMR (500 MHz, DMSO) δ 8.98 The combined organic layer was dried over Na2SO4, concen-
(s, 1H), 8.31 (d, J = 2.4 Hz, 1H), 8.07 (dd, J = 9.0, 2.5 Hz, 1H), trated under reduced pressure, and the crude compound was
7.76 (d, J = 9.0 Hz, 1H), 7.13 (s, 2H), 3.19 (t, J = 7.5 Hz, 2H), purified using silica gel column chromatography (10% MeOH/
1.89 (dd, J = 14.8, 7.4 Hz, 2H), 1.04 (t, J = 7.4 Hz, 3H), 0.36–0.33 CH2Cl2) to furnish compound 13b as a white solid (22 mg,
(m, 9H). 13C NMR (126 MHz, DMSO) δ 171.8, 152.7, 146.1, 87%). 1H NMR (400 MHz, CDCl3) δ 8.34 (d, J = 2.2 Hz, 1H),
144.4, 139.1, 138.2, 130.7, 128.9, 127.3, 121.3, 118.9, 115.3, 7.71 (d, J = 8.9 Hz, 1H), 7.37 (dd, J = 8.9, 2.4 Hz, 2H), 5.67 (d,
35.2, 22.8, 13.5, −0.9. MS (ESI) calculated for C18H22N6SSi, m/z J = 1.2 Hz, 2H), 3.13 (t, J = 7.4 Hz, 2H), 2.41 (t, J = 7.4 Hz, 2H),
382.14, found 383.14 (M + H)+.
1.94 (dd, J = 15.0, 7.5 Hz, 2H), 1.85–1.74 (m, 2H), 1.11–1.02 (m,
6H). 13C NMR (126 MHz, CDCl3) δ 172.0, 171.5, 150.9, 140.6,
138.2, 133.4, 126.8, 122.1, 120.2, 114.6, 39.8, 36.2, 23.3, 19.2,
13.9, 13.8. MS (ESI) calculated for C17H20N4OS, m/z 328.14,
Synthesis of compound 12g: 2-propyl-8-(1H-1,2,3-triazol-1-yl)-
thiazolo[4,5-c]quinolin-4-amine
To a stirred solution of compound 12f (20 mg, 0.0.52 mmol) in found 329.15 (M + H)+.
THF (1 mL) was added TBAF (20 mg, 0.078 mmol), and the
reaction mixture was stirred at room temperature for 10 h. The
reaction mixture was then diluted with water and extracted
Synthesis of compound 14: 8-bromo-2-propylthiazolo[4,5-c]-
quinolin-4-amine
with EtOAc (3 × 10 mL). The organic layer was dried over To a stirred solution of compound 8c (50 mg, 0.205 mmol) in
Na2SO4, concentrated under reduced pressure, and the crude CH3CN (1 mL) were added N-bromosuccinimide (44 mg,
material purified using silica gel column chromatography (5% 0.246 mmol), ammonium acetate (1.6 mg, 0.02 mmol) and the
MeOH/CH2Cl2) to obtain compound 12g as a white solid reaction mixture stirred at room temperature for 12 h. The
1
(14 mg, 87%). H NMR (500 MHz, DMSO) δ 8.94 (d, J = 1.1 Hz, reaction mixture was then diluted with water and extracted
1H), 8.29 (d, J = 2.4 Hz, 1H), 8.04 (dd, J = 9.0, 2.5 Hz, 1H), 7.98 with EtOAc (3 × 10 mL). The combined organic layer was dried
(d, J = 1.1 Hz, 1H), 7.75 (d, J = 9.0 Hz, 1H), 7.12 (s, 2H), 3.17 (t, over Na2SO4, concentrated under reduced pressure, and the
J = 7.5 Hz, 2H), 1.91–1.83 (m, 2H), 1.02 (t, J = 7.4 Hz, 3H). 13C crude material was purified using silica gel column chromato-
NMR (126 MHz, DMSO) δ 171.7, 152.8, 144.6, 139.1, 138.2, graphy (5% MeOH/CH2Cl2) to obtain compound 14 as a white
1192 | Org. Biomol. Chem., 2013, 11, 1179–1198
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1
solid (40 mg, 67%). H NMR (500 MHz, CDCl3) δ 7.86 (dd, J = Synthesis of compound 15d: N-(2-propylthiazolo[4,5-c]-
2.0, 0.6 Hz, 1H), 7.61 (dd, J = 3.9, 1.3 Hz, 2H), 5.68 (s, 2H), quinolin-4-yl)formamide
3.16–3.11 (m, 2H), 1.95 (dt, J = 15.0, 7.4 Hz, 2H), 1.09 (t, J = 7.4
To a stirred solution of compound 8c (50 mg, 0.205 mmol) in
1,4-dioxane (1 mL) were added 2,2,2-trifluoroethylformate
(60 μL, 0.616 mmol) and triethylamine (86 μL, 0.616 mmol).
The resulting mixture was stirred at 80 °C for 12 h. The reac-
Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 172.1, 151.8, 143.4,
139.3, 138.4, 132.1, 128.4, 126.9, 121.5, 115.9, 36.2, 23.4, 13.8.
MS (ESI) calculated for C13H12BrN3S, m/z 320.99, found 322.00
(M + H)+.
tion mixture was then diluted with water and extracted with
CH2Cl2 (3 × 10 mL). The combined organic layer was dried
over Na2SO4, concentrated under reduced pressure, and the
Synthesis of compound 15a: N-butyl-2-propylthiazolo[4,5-c]-
quinolin-4-amine
crude compound was purified using silica gel column chrom-
To a stirred solution of compound 8c (50 mg, 0.205 mmol) in
atography (3% MeOH/CH2Cl2) to furnish compound 15c as a
THF (1 mL) were added sodium hydride (10 mg, 0.41 mmol)
and 1-iodobutane (35 μL, 0.308 mmol), the resulting mixture
was stirred at room temperature for 6 h. The reaction mixture
was then diluted with water and extracted with CH2Cl2 (3 ×
10 mL). The organic layer was dried over Na2SO4, concentrated
under reduced pressure, and the crude compound was puri-
white solid (26 mg, 46%). 1H NMR (500 MHz, CDCl3) δ 9.84 (d,
J = 10.8 Hz, 1H), 8.86 (d, J = 10.3 Hz, 1H), 8.03–7.94 (m, 1H),
7.84 (ddd, J = 8.0, 1.4, 0.5 Hz, 1H), 7.67 (ddd, J = 8.5, 7.0, 1.4
Hz, 1H), 7.50 (ddd, J = 8.1, 7.1, 1.2 Hz, 1H), 3.24–3.08 (m, 2H),
2.05–1.87 (m, 2H), 1.09 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz,
CDCl3) δ 172.7, 162.0, 143.8, 143.2, 141.7, 137.5, 129.5, 128.8,
fied using silica gel column chromatography (5% MeOH/
125.8, 124.8, 121.9, 36.2, 23.1, 13.8. MS (ESI) calculated for
CH2Cl2) to obtain compound 15a (45 mg, 70%). 1H NMR
C14H13N3OS, m/z 271.08, found 272.09 (M + H)+.
(500 MHz, CDCl3) δ 7.81 (dd, J = 8.3, 0.5 Hz, 1H), 7.70–7.66 (m,
1H), 7.51 (ddd, J = 8.4, 7.1, 1.5 Hz, 1H), 7.23 (ddd, J = 8.0, 7.1,
1.1 Hz, 1H), 5.96 (t, J = 4.8 Hz, 1H), 3.77–3.69 (m, 2H),
Synthesis of compound 15e: N-(2-propylthiazolo[4,5-c]-
quinolin-4-yl)acetamide
3.15–3.08 (m, 2H), 1.99–1.89 (m, 2H), 1.75 (ddd, J = 14.9, 11.1,
To a stirred solution of compound 8c (20 mg, 0.082 mmol) in
7.5 Hz, 2H), 1.52 (dq, J = 14.7, 7.4 Hz, 2H), 1.08 (t, J = 7.4 Hz,
pyridine (1 mL) was added acetyl chloride (5.8 μL,
3H), 1.00 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ
0.082 mmol), the resulting mixture was stirred at room temp-
170.7, 151.2, 145.3, 138.9, 138.3, 128.6, 126.9, 124.5, 122.1,
erature for 1 h. The solvent was evaporated under reduced
119.3, 40.6, 36.0, 31.9, 23.3, 20.3, 14.0, 13.7. MS (ESI) calcu-
pressure and extracted with CH2Cl2 (3 × 10 mL). The combined
lated for C17H21N3S, m/z 299.15, found 300.14 (M + H)+.
organic layer was dried over Na2SO4, concentrated under
reduced pressure, and the crude compound was purified using
Compound 15b–c was synthesized similarly as compound
15a.
silica gel column chromatography (5% MeOH/CH2Cl2) to
furnish compound 15d as a white solid (18 mg, 78%). 1H NMR
15b: N-Hexyl-2-propylthiazolo[4,5-c]quinolin-4-amine
(500 MHz, CDCl3) δ 8.03 (d, J = 8.3 Hz, 1H), 7.88 (dd, J = 8.1,
White solid (9.9 mg, 72%, based on starting material recovery).
0.9 Hz, 1H), 7.72 (ddd, J = 8.4, 7.2, 1.3 Hz, 1H), 7.59–7.54 (m,
1H NMR (400 MHz, CDCl3) δ 7.81 (d, J = 8.3 Hz, 1H), 7.68 (dd, J
1H), 3.17 (td, J = 7.6, 2.3 Hz, 2H), 2.77 (s, 3H), 1.96 (dd, J =
= 7.9, 1.1 Hz, 1H), 7.51 (ddd, J = 8.5, 7.1, 1.5 Hz, 1H), 7.23
15.0, 7.5 Hz, 2H), 1.09 (t, J = 7.3 Hz, 3H). 13C NMR (126 MHz,
(ddd, J = 8.1, 7.2, 1.1 Hz, 1H), 5.96 (t, J = 4.6 Hz, 1H), 3.71 (dt, J
CDCl3) δ 173.0, 144.7, 138.3, 131.6, 130.0, 127.2, 126.5, 125.9,
= 7.2, 5.8 Hz, 2H), 3.11 (dd, J = 8.7, 6.5 Hz, 2H), 1.94 (dd, J =
125.3, 125.0, 121.2, 36.2, 25.7, 23.2, 13.8. MS (ESI) calculated
15.0, 7.5 Hz, 2H), 1.76 (t, J = 7.3 Hz, 2H), 1.53–1.44 (m, 2H),
1.36 (ddd, J = 6.9, 6.2, 3.3 Hz, 4H), 1.08 (t, J = 7.4 Hz, 3H), 0.91
for C15H15N3OS, m/z 285.09, found 286.11 (M + H)+.
Compounds 15f–15i were synthesized similarly as com-
(dd, J = 9.1, 5.1 Hz, 3H). 13C NMR (101 MHz, CDCl3) δ 170.9,
pound 15e.
151.3, 145.5, 139.1, 138.5, 128.8, 127.1, 124.6, 122.3, 119.5,
41.1, 36.2, 31.8, 29.9, 27.1, 23.5, 22.8, 14.2, 13.8. MS (ESI) cal-
15f: N-(2-Propylthiazolo[4,5-c]quinolin-4-yl)butyramide
White solid (20 mg, 78%). 1H NMR (500 MHz, CDCl3) δ 8.96 (s,
culated for C19H25N3S, m/z 327.18, found 328.18 (M + H)+.
15c: N-Hexadecyl-2-propylthiazolo[4,5-c]quinolin-4-amine
1H), 8.04 (d, J = 8.3 Hz, 1H), 7.82 (dd, J = 8.0, 1.0 Hz, 1H), 7.65
(ddd, J = 8.4, 7.1, 1.4 Hz, 1H), 7.49 (ddd, J = 8.1, 7.1, 1.1 Hz,
1H), 3.21–3.04 (m, 4H), 2.01–1.92 (m, 2H), 1.92–1.82 (m, 2H),
1.10 (td, J = 7.4, 6.1 Hz, 6H). 13C NMR (126 MHz, CDCl3) δ
172.1, 144.5, 143.5, 140.8, 138.6, 129.2, 129.2, 125.7, 124.6,
121.4, 39.9, 36.2, 23.2, 18.6, 14.1, 13.8. MS (ESI) calculated for
C17H19N3OS, m/z 313.12, found 314.13 (M + H)+.
White solid (26 mg, 68%). 1H NMR (400 MHz, CDCl3) δ 7.81
(d, J = 7.9 Hz, 1H), 7.68 (dd, J = 7.9, 1.1 Hz, 1H), 7.51 (ddd, J =
8.5, 7.1, 1.5 Hz, 1H), 7.23 (td, J = 7.1, 3.6 Hz, 1H), 5.96 (s, 1H),
3.71 (dd, J = 12.9, 7.1 Hz, 2H), 3.11 (dd, J = 8.7, 6.5 Hz, 2H),
1.98–1.88 (m, 2H), 1.76 (dt, J = 14.8, 7.4 Hz, 2H), 1.51–1.43 (m,
2H), 1.30–1.21 (m, 24H), 1.08 (t, J = 7.4 Hz, 3H), 0.88 (t, J = 6.8
Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 170.9, 151.4, 145.5,
139.1, 138.5, 128.7, 127.1, 124.6, 122.3, 119.5, 41.1, 36.8, 36.1,
15g: N-(2-Propylthiazolo[4,5-c]quinolin-4-yl)octanamide
32.1, 29.9, 29.8, 29.8, 29.8, 29.8, 29.8, 29.7, 29.6, 29.5, 27.3, White solid (25 mg, 83%). 1H NMR (400 MHz, CDCl3) δ 8.95 (s,
24.8, 23.5, 22.8, 14.3, 13.9. MS (ESI) calculated for C29H45N3S, 1H), 8.04 (d, J = 8.3 Hz, 1H), 7.82 (dd, J = 8.0, 0.9 Hz, 1H), 7.65
m/z 467.33, found 468.35 (M + H)+.
(ddd, J = 8.4, 7.1, 1.4 Hz, 1H), 7.52–7.44 (m, 1H), 3.15 (t, J = 7.5
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Hz, 4H), 1.96 (dd, J = 14.9, 7.4 Hz, 2H), 1.83 (dd, J = 15.2, 7.7 15k: 3-Azido-N-(2-propylthiazolo[4,5-c]quinolin-4-yl)propan-
Hz, 2H), 1.54–1.45 (m, 2H), 1.39–1.23 (m, 6H), 1.09 (t, J = 7.4 amide
Hz, 3H), 0.89 (t, J = 6.9 Hz, 3H). 13C NMR (126 MHz, CDCl3)
White solid (59 mg, 85%). 1H NMR (500 MHz, CDCl3) δ 9.07 (s,
δ 172.1, 144.5, 143.5, 140.8, 138.6, 129.3, 129.2, 125.7, 124.7,
1H), 8.03 (d, J = 8.3 Hz, 1H), 7.84 (dd, J = 8.0, 0.9 Hz, 1H), 7.67
121.4, 38.1, 36.2, 31.9, 29.8, 29.6, 29.3, 25.2, 23.2, 22.8, 14.2,
(ddd, J = 8.4, 7.1, 1.4 Hz, 1H), 7.53–7.48 (m, 1H), 3.82 (t, J = 6.6
13.8. MS (ESI) calculated for C21H27N3OS, m/z 369.19, found
Hz, 2H), 3.59 (s, 2H), 3.16 (t, J = 7.5 Hz, 2H), 2.01–1.91 (m, 2H),
370.22 (M + H)+.
1.09 (dd, J = 9.3, 5.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ
172.4, 144.1, 143.3, 141.1, 138.3, 129.4, 129.1, 125.9, 124.7,
121.5, 46.9, 37.6, 36.2, 23.2, 13.8. MS (ESI) calculated for
15h: N-(2-Propylthiazolo[4,5-c]quinolin-4-yl)palmitamide
C16H16N6OS, m/z 340.11, found 341.12 (M + H)+.
1
Yellow solid (33 mg, 84%). H NMR (500 MHz, CDCl3) δ 8.95
(s, 1H), 8.04 (d, J = 8.3 Hz, 1H), 7.83 (dd, J = 8.0, 1.0 Hz, 1H),
15l: N-(2-Propylthiazolo[4,5-c]quinolin-4-yl)pent-4-ynamide
7.65 (ddd, J = 8.4, 7.1, 1.4 Hz, 1H), 7.49 (ddd, J = 10.4, 5.8, 2.2
White solid (60 mg, 90%). 1H NMR (500 MHz, CDCl3) δ 9.02 (s,
Hz, 1H), 3.20–3.10 (m, 4H), 2.01–1.91 (m, 2H), 1.84 (dt, J =
1H), 8.05 (d, J = 8.3 Hz, 1H), 7.82 (dd, J = 8.0, 0.9 Hz, 1H), 7.66
(ddd, J = 8.4, 7.1, 1.4 Hz, 1H), 7.49 (ddd, J = 8.1, 7.1, 1.1 Hz,
1H), 3.49 (s, 2H), 3.15 (dd, J = 8.6, 6.5 Hz, 2H), 2.75 (ddd, J =
9.0, 6.9, 2.7 Hz, 2H), 2.03 (t, J = 2.7 Hz, 1H), 2.00–1.90 (m, 2H),
1.09 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 172.2,
144.2, 143.4, 141.0, 138.4, 129.3, 129.2, 125.8, 124.7, 121.5,
83.6, 68.9, 37.2, 36.2, 23.2, 14.3, 13.8. MS (ESI) calculated for
C18H17N3OS, m/z 323.11, found 324.13 (M + H)+.
15.2, 7.6 Hz, 2H), 1.53–1.44 (m, 2H), 1.43–1.34 (m, 2H),
1.34–1.19 (m, 20H), 1.09 (t, J = 7.4 Hz, 3H), 0.88 (t, J = 7.0 Hz,
3H). 13C NMR (126 MHz, CDCl3) δ 172.1, 144.5, 143.6, 140.8,
138.6, 129.3, 129.2, 125.7, 124.7, 121.5, 38.1, 36.2, 32.1, 29.8,
29.8, 29.8, 29.7, 29.6, 29.6, 29.5, 25.2, 23.2, 22.8, 14.3, 13.8. MS
(ESI) calculated for C29H43N3OS, m/z 481.31, found 482.33 (M
+ H)+.
15i: N-(2-Propylthiazolo[4,5-c]quinolin-4-yl)benzamide
Synthesis of compound 15m: methyl (2-propylthiazolo[4,5-c]-
White solid (24 mg, 96%). 1H NMR (400 MHz, CDCl3) δ 9.70 (s,
1H), 8.24 (d, J = 8.5 Hz, 1H), 8.07 (d, J = 7.4 Hz, 2H), 7.86 (d, J =
7.6 Hz, 1H), 7.69 (t, J = 7.5 Hz, 1H), 7.62–7.51 (m, 4H), 3.19 (t,
J = 7.6 Hz, 2H), 1.98 (dt, J = 14.9, 7.5 Hz, 2H), 1.12 (t, J = 7.4 Hz,
3H). 13C NMR (126 MHz, CDCl3) δ 172.4, 164.7, 144.7, 143.8,
140.9, 139.3, 135.0, 132.4, 130.1, 129.2, 129.0, 127.7, 126.1,
124.5, 121.8, 36.2, 23.4, 13.9. MS (ESI) calculated for
C20H17N3OS, m/z 347.11, found 348.12 (M + H)+.
quinolin-4-yl)carbamate
To a stirred solution of 8c (50 mg, 0.205 mmol) in CH2Cl2
(2 mL) were added triethylamine (57 μL, 0.410 mmol) and
methyl chloroformate (32 μL, 0.3075 mmol), and the resulting
mixture was stirred at room temperature for 5 h. The reaction
mixture was diluted with water and extracted with CH2Cl2 (3 ×
10 mL). The organic layer was dried over Na2SO4, concentrated
under reduced pressure, and the crude residue was purified by
silica gel column chromatography (5% MeOH/CH2Cl2) to
obtain compound 15m (58 mg, 94%) as a white solid. 1H NMR
(500 MHz, CDCl3) δ 8.51 (s, 1H), 8.16 (d, J = 8.4 Hz, 1H), 7.82
(dd, J = 8.0, 0.9 Hz, 1H), 7.69–7.63 (m, 1H), 7.52–7.46 (m, 1H),
3.91 (s, 3H), 3.17–3.11 (m, 2H), 1.95 (dt, J = 14.8, 7.4 Hz, 2H),
1.09 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 172.2,
152.2, 144.3, 143.8, 140.8, 138.6, 129.7, 129.2, 125.7, 124.5,
121.4, 52.9, 36.2, 23.3, 13.8. MS (ESI) calculated for
C15H15N3O2S, m/z 301.09, found 302.17 (M + H)+.
Synthesis of compound 15j: 2-azido-N-(2-propylthiazolo[4,5-c]-
quinolin-4-yl)acetamide
To
a stirred solution of 2-azidoacetic acid (18.6 mg,
0.123 mmol) in DMF (1 mL) were added triethylamine (23 μL,
0.164 mmol) and HBTU (46.6 mg, 0.123 mmol), and the result-
ing mixture was stirred at room temperature for 15 min. Com-
pound 8c (20 mg, 0.082 mmol) was then added, and stirring
continued for 4 h. The reaction mixture was diluted with water
and extracted with CH2Cl2 (3 × 10 mL). The organic layer was
dried over Na2SO4, concentrated under reduced pressure, and
the crude residue was purified by silica gel column chromato-
graphy (5% MeOH/CH2Cl2) to obtain compound 15j as a white
Compounds 15n–15p were synthesized similarly as com-
pound 15m.
Synthesis of compound 15n: ethyl (2-propylthiazolo[4,5-c]
quinolin-4-yl)carbamate
1
solid (50 mg, 96%). H NMR (500 MHz, CDCl3) δ 9.25 (d, J =
1
0.6 Hz, 1H), 8.03 (d, J = 8.1 Hz, 1H), 7.84 (dd, J = 8.0, 1.0 Hz, Yellow solid (50 mg, 77%). H NMR (500 MHz, CDCl3) δ 8.48
1H), 7.68 (ddd, J = 8.4, 7.1, 1.4 Hz, 1H), 7.52 (ddd, J = 8.1, 7.2, (s, 1H), 8.17 (t, J = 8.3 Hz, 1H), 8.17 (t, J = 8.3 Hz, 1H), 7.81 (dd,
1.1 Hz, 1H), 4.86 (s, 2H), 3.17 (t, J = 7.5 Hz, 2H), 2.01–1.92 (m, J = 8.0, 1.0 Hz, 1H), 7.81 (dd, J = 8.0, 1.0 Hz, 1H), 7.69–7.61 (m,
2H), 1.10 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) 1H), 7.51–7.45 (m, 1H), 4.36 (q, J = 7.1 Hz, 2H), 3.18–3.12 (m,
δ 172.7, 143.6, 143.1, 141.4, 138.2, 129.5, 129.1, 126.2, 124.8, 2H), 2.01–1.90 (m, 2H), 1.39 (t, J = 7.1 Hz, 3H), 1.09 (t, J = 7.4
121.6, 54.7, 36.2, 23.2, 13.8. MS (ESI) calculated for Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 172.1, 151.6, 144.5,
C15H14N6OS, m/z 326.10, found 327.11 (M + H)+.
Compounds 15k–15l were synthesized similarly as com- 23.3, 14.7, 13.8. MS (ESI) calculated for C16H17N3O2S, m/z
pound 15j.
315.10, found 316.19 (M + H)+.
143.8, 140.8, 138.6, 129.8, 129.1, 125.7, 124.5, 121.3, 61.8, 36.2,
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Synthesis of compound 15o: butyl (2-propylthiazolo[4,5-c]-
quinolin-4-yl)carbamate
layer was dried over Na2SO4, concentrated under reduced
pressure, and the crude residue was purified by silica gel
column chromatography (5% MeOH/CH2Cl2) to obtain com-
White solid (52 mg, 74%). 1H NMR (500 MHz, CDCl3) δ 8.47 (s,
1H), 8.18–8.13 (m, 1H), 7.81 (dd, J = 8.0, 0.9 Hz, 1H), 7.66 (qd,
J = 8.5, 2.6 Hz, 1H), 7.52–7.45 (m, 1H), 4.31 (t, J = 6.7 Hz, 2H),
3.19–3.11 (m, 2H), 2.02–1.91 (m, 2H), 1.80–1.70 (m, 2H),
1.54–1.44 (m, 2H), 1.13–1.06 (m, 3H), 0.98 (t, J = 7.4 Hz, 3H).
13C NMR (126 MHz, CDCl3) δ 172.1, 151.8, 144.5, 143.7, 140.8,
138.6, 129.8, 129.1, 125.7, 124.5, 121.3, 65.7, 36.2, 31.0, 23.3,
19.2, 13.9, 13.8. MS (ESI) calculated for C18H21N3O2S, m/z
343.14, found 344.14 (M + H)+.
1
pound 15r as a white solid (34 mg, 65%). H NMR (500 MHz,
CDCl3) δ 11.91 (s, 1H), 7.73 (d, J = 7.8 Hz, 1H), 7.58 (t, J = 7.4
Hz, 1H), 7.40 (t, J = 7.6 Hz, 2H), 3.29 (s, 3H), 3.20–3.15 (m, 2H),
1.97–1.87 (m, 2H), 1.07 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz,
CDCl3) δ 173.0, 148.8, 143.4, 141.6, 133.5, 130.6, 125.4, 125.2,
117.5, 116.9, 43.2, 36.3, 23.7, 13.8. MS (ESI) calculated for
C14H15N3O2S2, m/z 321.06, found 322.06 (M + H)+.
Compounds 15s–15v were synthesized similarly as com-
pound 15r.
Synthesis of compound 15p: octyl (2-propylthiazolo[4,5-c]-
quinolin-4-yl)carbamate
Synthesis of compound 15s: N-(2-propylthiazolo[4,5-c]-
quinolin-4-yl)ethanesulfonamide
White solid (60 mg, 73%). 1H NMR (500 MHz, CDCl3) δ 8.47 (s,
1H), 8.17–8.13 (m, 1H), 7.81 (dd, J = 8.0, 1.0 Hz, 1H), 7.68–7.61
(m, 1H), 7.47 (ddd, J = 8.1, 7.2, 1.1 Hz, 1H), 4.30 (t, J = 6.7 Hz,
2H), 3.15 (dd, J = 9.7, 5.5 Hz, 2H), 2.02–1.90 (m, 3H), 1.80–1.71
(m, 2H), 1.51–1.39 (m, 3H), 1.38–1.22 (m, 10H), 1.09 (dd, J =
9.0, 5.8 Hz, 5H), 0.88 (t, J = 7.0 Hz, 3H). 13C NMR (126 MHz,
CDCl3) δ 172.1, 151.8, 144.5, 143.9, 140.8, 138.6, 129.7, 129.1,
125.6, 124.5, 121.3, 66.0, 36.2, 31.9, 29.4, 29.4, 29.0, 26.0, 23.3,
22.8, 14.2, 13.8. MS (ESI) calculated for C22H29N3O2S, m/z
399.20, found 400.21 (M + H)+.
1
White solid (35 mg, 63%). H NMR (500 MHz, CDCl3) δ 12.05
(s, 1H), 7.72 (d, J = 6.7 Hz, 1H), 7.58 (t, J = 7.1 Hz, 1H), 7.40 (d,
J = 7.3 Hz, 2H), 3.34 (d, J = 5.0 Hz, 2H), 3.18 (t, J = 7.7 Hz, 2H),
1.99–1.87 (m, 2H), 1.46 (t, J = 7.4 Hz, 3H), 1.07 (t, J = 7.4 Hz,
3H). 13C NMR (126 MHz, CDCl3) δ 172.9, 149.9, 143.3, 141.9,
133.5, 130.7, 125.5, 125.0, 117.7, 116.8, 49.4, 36.3, 23.8, 13.8,
8.3. MS (ESI) calculated for C15H17N3O2S2, m/z 335.08, found
336.08 (M + H)+.
Synthesis of compound 15t: N-(2-propylthiazolo[4,5-c]quinolin-
4-yl)propane-1-sulfonamide
Synthesis of compound 15q: 1-(2-propylthiazolo[4,5-c]-
quinolin-4-yl)urea
1
White solid (35 mg, 61%). H NMR (500 MHz, CDCl3) δ 12.05
(s, 1H), 7.72 (d, J = 6.7 Hz, 1H), 7.58 (t, J = 7.1 Hz, 1H), 7.40 (d,
J = 7.3 Hz, 2H), 3.34 (d, J = 5.0 Hz, 2H), 3.18 (t, J = 7.7 Hz, 2H),
1.99–1.87 (m, 2H), 1.46 (t, J = 7.4 Hz, 3H), 1.07 (t, J = 7.4 Hz,
3H). 13C NMR (126 MHz, CDCl3) δ 172.9, 149.9, 143.3, 141.9,
133.5, 130.7, 125.5, 125.0, 117.7, 116.8, 49.4, 36.3, 23.8, 13.8,
8.3. MS (ESI) calculated for C16H19N3O2S2, m/z 349.09, found
350.09 (M + H)+.
To a stirred solution of 8c (50 mg, 0.205 mmol) in acetonitrile
(2 mL) were added NaHCO3 (26 mg, 0.307 mmol) and chloro-
sulfonyl isocyanate (27 μL, 0.307 mmol), and the resulting
mixture was stirred at room temperature for 1 h. Then, one
more equivalent of chlorosulfonyl isocyanate (27 μL,
0.307 mmol) was added and the reaction mixture was stirred at
room temperature for additional 1 h. The reaction mixture was
diluted with water and extracted with EtOAc (3 × 10 mL). The
organic layer was dried over Na2SO4, concentrated under
reduced pressure, and the crude residue was purified by silica
gel column chromatography (5% MeOH/CH2Cl2) to obtain
compound 15q as a white solid (26 mg, 45% starting material
Synthesis of compound 15u: N-(2-propylthiazolo[4,5-c]-
quinolin-4-yl)butane-1-sulfonamide
1
White solid (37 mg, 62%). H NMR (500 MHz, CDCl3) δ 12.04
(s, 1H), 7.71 (d, J = 6.7 Hz, 1H), 7.57 (t, J = 7.0 Hz, 1H), 7.39 (d,
J = 6.2 Hz, 2H), 3.32 (s, 2H), 3.18 (t, J = 7.7 Hz, 2H), 2.00–1.82
(m, 4H), 1.55–1.40 (m, 2H), 1.07 (t, J = 7.4 Hz, 3H), 0.94 (t, J =
7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 172.8, 149.7, 143.3,
141.9, 133.5, 130.7, 125.5, 125.0, 117.6, 116.8, 54.8, 36.3, 25.5,
23.8, 21.7, 13.8, 13.8. MS (ESI) calculated for C17H21N3O2S2, m/
z 363.11, found 364.11 (M + H)+.
1
was recovered). H NMR (500 MHz, CDCl3) δ 9.84 (d, J = 10.8
Hz, 1H), 8.86 (d, J = 10.3 Hz, 1H), 8.03–7.94 (m, 1H), 7.84 (ddd,
J = 8.0, 1.4, 0.5 Hz, 1H), 7.67 (ddd, J = 8.5, 7.0, 1.4 Hz, 1H), 7.50
(ddd, J = 8.1, 7.1, 1.2 Hz, 1H), 3.24–3.08 (m, 2H), 2.05–1.87 (m,
2H), 1.09 (t, J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ
172.7, 162.0, 143.8, 143.2, 141.7, 137.5, 129.5, 128.8, 125.8,
124.8, 121.9, 36.2, 23.1, 13.8. MS (ESI) calculated for
C14H14N4OS, m/z 286.09, found 287.09 (M + H)+.
Synthesis of compound 15v: 4-methyl-N-(2-propylthiazolo[4,5-c]-
quinolin-4-yl)benzenesulfonamide
1
White solid (40 mg, 61%). H NMR (500 MHz, CDCl3) δ 12.04
(s, 1H), 7.71 (d, J = 6.7 Hz, 1H), 7.57 (t, J = 7.0 Hz, 1H), 7.39 (d,
J = 6.2 Hz, 2H), 3.32 (s, 2H), 3.18 (t, J = 7.7 Hz, 2H), 2.00–1.82
Synthesis of compound 15r: N-(2-propylthiazolo[4,5-c]-
quinolin-4-yl)methanesulfonamide
To a stirred solution of 8c (40 mg, 0.146 mmol) in dichloro- (m, 4H), 1.55–1.40 (m, 2H), 1.07 (t, J = 7.4 Hz, 3H), 0.94 (t, J =
methane (2 mL) was added methyl methanesulfonyl chloride 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 172.8, 149.0, 143.4,
(26 μL, 0.292 mmol), and the resulting mixture was stirred at 143.0, 142.1, 139.9, 133.5, 130.7, 129.4, 126.7, 125.5, 125.1,
room temperature for 12 h. The reaction mixture was diluted 117.7, 117.0, 36.2, 23.67, 21.7, 13.8. MS (ESI) calculated for
with water and extracted with CH2Cl2 (3 × 10 mL). The organic C20H19N3O2S2, m/z 397.09, found 398.10 (M + H)+.
This journal is © The Royal Society of Chemistry 2013
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Organic & Biomolecular Chemistry
Synthesis of compound 15w: diethyl (2-propylthiazolo[4,5-c]
quinolin-4-yl)phosphoramidate
2H), 1.91 (dt, J = 15.2, 7.6 Hz, 2H), 1.55–1.45 (m, 2H), 1.01 (t,
J = 7.4 Hz, 3H). 13C NMR (126 MHz, CDCl3) δ 172.8, 161.9,
143.7, 143.2, 141.6, 137.4, 129.4, 128.7, 125.7, 124.7, 121.8,
33.8, 31.6, 22.2, 13.8. MS (ESI) calculated for C15H15N3OS, m/z
285.09, found 286.10 (M + H)+.
To a stirred solution of 8c (50 mg, 0.205 mmol) in CH2Cl2
(2 mL) was added diethyl chlorophosphate (60 μL,
0.810 mmol), and the resulting mixture was stirred at room
temperature for 12 h. The reaction mixture was diluted with
water and extracted with CH2Cl2 (3 × 10 mL). The organic layer
was dried over Na2SO4, concentrated under reduced pressure,
and the crude residue was purified by silica gel column chrom-
atography (5% MeOH/CH2Cl2) to obtain compound 15w as a
white solid (45 mg, 56%). 1H NMR (500 MHz, DMSO) δ 8.41 (d,
J = 10.1 Hz, 1H), 7.96 (d, J = 7.8 Hz, 1H), 7.83 (d, J = 8.3 Hz,
1H), 7.68 (dd, J = 11.2, 4.0 Hz, 1H), 7.49 (t, J = 7.4 Hz, 1H), 4.24
(p, J = 7.2 Hz, 4H), 3.20 (t, J = 7.5 Hz, 2H), 1.94–1.85 (m, 2H),
1.27 (t, J = 7.0 Hz, 6H), 1.02 (t, J = 7.4 Hz, 3H). 13C NMR
(126 MHz, DMSO) δ 171.9, 147.0, 143.0, 140.3, 138.0 (d, J =
10.4 Hz), 131.2, 129.2 (d, J = 207.2 Hz), 127.8 (d, J = 94.8 Hz),
124.9 (d, J = 12.8 Hz), 120.2, 63.1 (d, J = 5.6 Hz), 35.2, 63.1 (d,
J = 5.6 Hz), 16.2 (d, J = 6.8 Hz) 13.5. MS (ESI) calculated for
C17H22N3O3PS, m/z 379.11, found 380.12 (M + H)+.
Compounds 18b–c were synthesized similarly as compound
15d.
Synthesis of compound 18b: N-(2-butylthiazolo[4,5-c]quinolin-
4-yl)acetamide
White solid (50 mg, 85%). 1H NMR (500 MHz, CDCl3) δ 8.97 (s,
1H), 8.02 (d, J = 8.3 Hz, 1H), 7.82 (ddd, J = 8.0, 1.4, 0.5 Hz, 1H),
7.65 (ddd, J = 8.4, 7.1, 1.4 Hz, 1H), 7.49 (ddd, J = 8.1, 7.1, 1.2
Hz, 1H), 3.21–3.13 (m, 2H), 2.82 (s, 3H), 1.96–1.85 (m, 2H),
1.55–1.44 (m, 2H), 1.04–0.96 (m, 3H). 13C NMR (126 MHz,
CDCl3) δ 172.4, 144.4, 143.4, 140.9, 138.5, 129.2, 129.1, 125.7,
124.7, 121.5, 34.0, 31.8, 25.9, 22.3, 13.9. MS (ESI) calculated for
C16H17N3OS, m/z 299.11, found 300.11 (M + H)+.
Synthesis of compound 18c: N-(2-butylthiazolo[4,5-c]quinolin-
4-yl)butyramide
White solid (55 mg, 87%). 1H NMR (500 MHz, CDCl3) δ 8.95 (s,
1H), 8.05 (d, J = 8.3 Hz, 1H), 7.82 (ddd, J = 8.0, 1.4, 0.4 Hz, 1H),
7.65 (ddd, J = 8.4, 7.1, 1.4 Hz, 1H), 7.49 (ddd, J = 8.1, 7.1, 1.2
Hz, 1H), 3.22–3.06 (m, 4H), 1.95–1.83 (m, 4H), 1.54–1.45 (m,
2H), 1.10 (t, J = 7.4 Hz, 3H), 1.00 (t, J = 7.4 Hz, 3H). 13C NMR
(126 MHz, CDCl3) δ 172.2, 144.3, 143.4, 140.7, 138.4, 129.1,
129.0, 125.6, 124.5, 121.3, 39.8, 33.8, 31.7, 22.2, 18.5, 14.0,
13.8. MS (ESI) calculated for C18H21N3OS, m/z 327.14, found
328.14 (M + H)+.
Synthesis of compound 17: 1-(2-propylthiazolo[4,5-c]quinolin-
4-yl)guanidine
Compound 7c (100 mg, 0.411 mmol) was dissolved in POCl3
(3 mL) and stirred at 100 °C for 1 h. POCl3 was evaporated
under reduced pressure, ice cold water was added and the
residue was extracted with CH2Cl2 (3 × 10 mL). The organic
layer was dried over Na2SO4, concentrated under reduced
pressure to give crude 16 (100 mg, 93%) which was used
further without purification. To a stirred solution of guanidine
(62 mg, 0.615 mmol) in 1,4 dioxane (2 mL) was added NaH Human TLR-7/-8 reporter gene assays (NF-κB induction)
(12 mg, 0.492 mmol), the reaction mixtures was stirred at
60 °C for 30 min. Compound 16 (30 mg, 0.123 mmol) in DMF
The induction of NF-κB was quantified using HEK-Blue-7
(hTLR7-specific) and HEK-Blue-8 (hTLR8-specific) cells as pre-
(2 mL) was added and heating was continued at 90 °C for 12 h.
viously described by us.27,32,33 HEK293 cells stably co-trans-
The reaction mixture was diluted with water and extracted with
fected with human TLR7 or human TLR8 and secreted
CH2Cl2 (3 × 10 mL). The organic layer was dried over Na2SO4,
alkaline phosphatase (sAP), were maintained in HEK-Blue™
concentrated under reduced pressure, and the crude residue
Selection medium containing zeocin and normocin. Stable
was purified by silica gel column chromatography (10%
expression of secreted alkaline phosphatase (sAP) under
MeOH/CH2Cl2) to obtain compound 17 as a yellow solid
control of NF-κB/AP-1 promoters is inducible by appropriate
1
(25 mg, 71%). H NMR (500 MHz, MeOD) δ 8.05 (dd, J = 8.4,
TLR agonists, and extracellular sAP in the supernatant is pro-
0.5 Hz, 1H), 8.00–7.96 (m, 1H), 7.75 (ddd, J = 8.4, 7.1, 1.4 Hz,
portional to NF-κB induction. HEK-Blue cells were incubated
1H), 7.62 (ddd, J = 8.2, 7.1, 1.2 Hz, 1H), 3.25 (dd, J = 8.7, 6.4
at a density of ∼105 cells per ml in a volume of 80 μL per well,
Hz, 2H), 2.06–1.97 (m, 2H), 1.11 (t, J = 7.4 Hz, 3H). 13C NMR
in 384-well, flat-bottomed, cell culture-treated microtiter plates
until confluency was achieved, and subsequently stimulated
with graded concentrations of stimuli. sAP was assayed spec-
trophotometrically using an alkaline phosphatase-specific
chromogen (present in HEK-detection medium as supplied by
the vendor) at 620 nm.
(126 MHz, MeOD) δ 175.2, 157.5, 146.0, 143.5, 142.7, 138.9,
131.0, 129.3, 128.0, 125.9, 122.5, 36.9, 24.2, 14.0. MS (ESI) cal-
culated for C14H15N5S, m/z 285.10, found 286.11 (M + H)+.
Compounds 18a were synthesized similarly as compound
15c.
Synthesis of compound 18a: N-(2-butylthiazolo[4,5-c]quinolin-
Immunoassays for Interferon (IFN)-α, IFN-γ, Interleukin (IL)-
4-yl)formamide
12, IL-18, and cytokines
White solid (32 mg, 71%). 1H NMR (500 MHz, CDCl3) δ 9.85 Fresh human peripheral blood mononuclear cells (hPBMC)
(d, J = 10.8 Hz, 1H), 8.86 (d, J = 10.3 Hz, 1H), 7.99 (d, J = 8.4 were isolated from human blood obtained by venipuncture
Hz, 1H), 7.85 (dd, J = 8.0, 0.9 Hz, 1H), 7.67 (ddd, J = 8.4, 7.1, with informed consent and as per institutional guidelines on
1.4 Hz, 1H), 7.50 (ddd, J = 8.1, 7.1, 1.1 Hz, 1H), 3.20–3.16 (m, Ficoll-Hypaque gradients as described elsewhere.62 Aliquots of
1196 | Org. Biomol. Chem., 2013, 11, 1179–1198
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Paper
PBMCs (105 cells in 100 μL per well) were stimulated for 12 h 14 T. Kaisho and S. Akira, Biochim. Biophys. Acta, 2002,
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Rabbit immunization and antigen-specific ELISA
All experiments were performed at Harlan Laboratories (India-
napolis, IN) in accordance with institutional guidelines (Uni-
versity of Kansas IACUC permit # 119-06). Cohorts of adult
female New Zealand White rabbits (n = 3 per cohort) were
immunized intramuscularly in the flank region with (a) 100 μg
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Acknowledgements
This work was supported by NIH/NIAID contract
HHSN272200900033C.
30 D. B. Salunke, N. M. Shukla, E. Yoo, B. M. Crall,
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