Discovery of potent BACE-1 inhibitors containing a new hydroxyethylene (HE) Scaffold: Exploration of P1′ alkoxy residues and an aminoethylene (AE) central core

Article abstract of DOI:10.1016/j.bmc.2009.12.051

In a preceding study we have described the development of a new hydroxyethylene (HE) core motif displaying P1 aryloxymethyl and P1′ methoxy substituents delivering potent BACE-1 inhibitors. In a continuation of this work we have now explored the SAR of the S1′ pocket by introducing a set of P1′ alkoxy groups and evaluated them as BACE-1 inhibitors. Previously the P1 and P1′ positions of the classical HE template have been relatively little explored due to the complexity of the chemical routes involved in modifications at these positions. However, the chemistries developed for the current HE template renders substituents in both the P1 and P1′ positions readily available for SAR exploration. The BACE-1 inhibitors prepared displayed K_i values in the range of 1-20 nM, where the most potent compounds featured small P1′ groups. The cathepsin D selectivity which was high for the smallest P1′ substituents (P1′ = ethoxy, fold selectively >1500) dropped for larger groups (P1′ = benzyloxy, fold selectivity of 3). We have also confirmed the importance of both the hydroxyl group and its stereochemistry preference for this HE transition state isostere by preparing both the deoxygenated analogue and by inverting the configuration of the hydroxyl group to the R-configuration, which as expected resulted in large activity drops. Finally substituting the hydroxyl group by an amino group having the same configuration (S), which previously have been described to deliver potent BACE-1 inhibitors with advantageous properties, surprisingly resulted in a large drop in the inhibitory activity.

Full text of DOI:10.1016/j.bmc.2009.12.051

Bioorganic & Medicinal Chemistry 18 (2010) 1711–1723
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Discovery of potent BACE-1 inhibitors containing a new hydroxyethylene
(HE) Scaffold: Exploration of P10 alkoxy residues and an aminoethylene
(AE) central core
Catarina Björklund a, Hans Adolfsson a, Katarina Jansson b, Jimmy Lindberg b, Lotta Vrang b,
Anders Hallberg c, Åsa Rosenquist b, Bertil Samuelsson a,b,
*
a Department of Organic Chemistry, Arrhenius Laboratory, Stockholm University, SE-106 91 Stockholm, Sweden
b Medivir AB, PO Box 1086, SE-141 22 Huddinge, Sweden
c Department of Medicinal Chemistry, BMC, Uppsala University, Box 574, SE-751 23 Uppsala, Sweden
a r t i c l e i n f o
a b s t r a c t
Article history:
In a preceding study we have described the development of a new hydroxyethylene (HE) core motif dis-
playing P1 aryloxymethyl and P10 methoxy substituents delivering potent BACE-1 inhibitors. In a contin-
uation of this work we have now explored the SAR of the S10 pocket by introducing a set of P10 alkoxy
groups and evaluated them as BACE-1 inhibitors. Previously the P1 and P10 positions of the classical
HE template have been relatively little explored due to the complexity of the chemical routes involved
in modifications at these positions. However, the chemistries developed for the current HE template ren-
ders substituents in both the P1 and P10 positions readily available for SAR exploration. The BACE-1 inhib-
itors prepared displayed Ki values in the range of 1–20 nM, where the most potent compounds featured
small P10 groups. The cathepsin D selectivity which was high for the smallest P10 substituents (P10 = eth-
oxy, fold selectively >1500) dropped for larger groups (P10 = benzyloxy, fold selectivity of 3). We have also
confirmed the importance of both the hydroxyl group and its stereochemistry preference for this HE tran-
sition state isostere by preparing both the deoxygenated analogue and by inverting the configuration of
the hydroxyl group to the R-configuration, which as expected resulted in large activity drops. Finally
substituting the hydroxyl group by an amino group having the same configuration (S), which previously
have been described to deliver potent BACE-1 inhibitors with advantageous properties, surprisingly
resulted in a large drop in the inhibitory activity.
Received 6 August 2009
Revised 23 November 2009
Accepted 20 December 2009
Available online 11 January 2010
Keywords:
Alzhemier’s disease
BACE-1 inhibitors
Hydroxyethylene (HE) isostere
Aminoethylene (AE) isostere
P10 modifications
Peptidomimetics
Ó 2010 Elsevier Ltd. All rights reserved.
1. Introduction
In a preceding study from our group we have disclosed the
development of a novel HE transition state isostere, where a di-flu-
Alzheimer’s disease (AD) is a devastating neurodegenerative
disorder characterized by the progressive formation of insoluble
amyloid plaques and neurofibrillary tangles in the brain.1,2 The ma-
jor component of the plaques, amyloid b peptides (Ab) is generated
from amyloid precursor protein (APP) by b-site amyloid precursor
oro phenoxymethyl group was introduced in the P1 position and a
methoxy group in the P10 position furnishing highly potent inhib-
itors of BACE-1 (i.e., lead compound 1),11 which moreover exhibits
very promising selectivity over cathepsin D. While BACE-1 HE isos-
teres generally employ a methyl group in the P10 position we rea-
soned based on modeling and X-ray crystallography studies that
both BACE-1 potency and cathepsin D selectivity might be en-
hanced by replacing this methyl group in the P10 position by a
more polar but still small methoxy group, which we were pleased
to note was also the case. As deduced from 3D structural data11,12
the oxygen of the methoxy group is involved in hydrogen bonding
to a structural water molecule which in turn is hydrogen bonded to
Thr72 of the BACE-1 enzyme. In contrast, cathepsin D has a lipo-
philic S10 pocket and a polar methoxy group is subsequently less
well accommodated than a methyl group, resulting in reduced
cathepsin D potency and the desired enhanced selectivity over
cathepsin D. The natural BACE-1 substrate has an Asp residue in
protein (APP)-cleaving enzyme 1 (BACE-1) and c-secretase-medi-
ated stepwise cleavages.3 It is hypothesized that accumulation of
these Ab peptides and subsequent formation of plaques in the
brain may be responsible for the onset and progression of the dis-
ease where the neurotoxicity of Ab peptides cause neuronal cell
death and brain inflammation ultimately leading to the dementia
and AD.4,5 As a result of the amyloid hypothesis, BACE-1 has been
vigorously pursued as a prime molecular target for therapeutic
intervention in AD.6–10
* Corresponding author. Tel.: +46 8 6083104; fax: +46 8 6083199.
E-mail address: bertil.samuelsson@medivir.com (B. Samuelsson).
0968-0896/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2009.12.051

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C. Björklund et al. / Bioorg. Med. Chem. 18 (2010) 1711–1723
the P10 position13 and it has previously been reported14 that selec-
tivity over cathepsin D can be enhanced by increasing the polarity
of the side chain at the P10 position although the BACE-1 inhibitors
Scheme 1. Glycoside 3 was treated with Ag2O and reagent 12a or
12b from chemset 12 (Fig. 2) in DMF at room temperature to fur-
nish compounds 4a and 4b in 47% and 34% yield, respectively.23
Previous experience in our lab prompted us to use this method
but due to low or zero yields in the following alkylation attempts,
a switch to NaH was made with improved results. To prepare 4c–e,
glycoside 3 was treated with NaH in DMF at 0 °C and then reacted
with the reagents 12c–e from chemset 12 in DMF to deliver 4c–e
in 19%, 49% and 23% yield, respectively. Hydrolysis of the methyl
glycosides 4a–e in refluxing 1,4-dioxane in the presence of sulfuric
acid for ꢀ1 h followed by quenching with aqueous sodium carbon-
ate afforded the desired products 5a–e in 47%, 82%, 80%, 55% and
67% yield, respectively.24 Subsequent oxidation of the anomeric
hydroxyl group using pyridinium dichromate in dichloromethane
(DCM) provided the corresponding lactones 6a–e in 96%, 68%,
81%, 38% and 83% yield, respectively.25 These lactones 6a–e were
ring opened with (S)-2-amino-N-benzyl-3-methyl-butyramide
upon heating in diisopropylethylamine (DIPEA) using 2-hydroxy-
pyridine as an activator of the lactone to give the ring opened
amides 7a–e in 70%, 84%, 87%, 77% and 97% yield, respectively.26,27
The amine, (S)-2-amino-N-benzyl-3-methyl-butyramide was syn-
thesised from Boc-Val-OH and benzylamine.11 Reduction of the
azide group in compounds 7a–e was achieved using triphenyl-
phosphine (Ph3P) in methanol containing a few drops of water pro-
viding the corresponding amines which were coupled without
prior purification with 5-(methanesulfonyl-methyl-amino)-N0-
(1-phenyl-ethyl)-isophthalacid17 using benzotriazole-1-yloxytris-
(pyrrolidino)phosphonium hexafluorphosphate (Py-BOP) and DI-
PEA in DCM furnishing the target compounds 2a–d and 2f in
64%, 67%, 97%, 58%, and 43% yield, respectively.21,28 Compound
7d was reacted with hydrogen over a catalytically amount of pal-
ladium on activated carbon to reduce both the azide group and
the double bond of the P10 allyl group followed by coupling with
the P2–P3 group using the same protocol, vide supra, to give prod-
uct 2e in 88% yield.
explored all displayed BACE-1 Ki values at best in the low lM.
Based on the encouraging results obtained in our previous
study, cf. inhibitor 1, we have now studied a range of P10 alkoxy
groups, to explore the selectivity differences between the S10 pock-
ets of BACE-1 and cathepsin D,14 respectively. A number of P10
groups, all featuring the alkoxy motif, have been synthesized and
evaluated as BACE-1 inhibitors where the chemistry developed
for this novel HE template has allowed the facile introduction of
substituents in both the P1 and P10 positions. Previously these
positions have been relatively little explored due to the complexity
of the chemical synthesis involved in modifications of the classical
HE template.15 Several groups have reported on BACE-1 inhibitors
containing substituted isophthalamides as highly promising P2–P3
groups.16 All our inhibitors in this series are equipped with the 5-
substituted isophthalamide reported by Stachel et al. (Fig. 1).17
We herein report a series of new inhibitors, among those 2a–2f,
displayed BACE-1 Ki values in the range 1–20 nM, with the most
potent compounds being those inhibitors having the smallest alk-
oxy P10 groups and where the caphepsin D selectivity was high for
the smallest P10 substituents. Furthermore we confirmed the
importance of a hydroxyl group with S-configuration and that a
replacement of this hydroxyl group for an amine, unexpectedly
was non-productive in this series.
2. Results and discussion
2.1. Chemistry
The target compounds 2a–j were prepared as outlined in
Schemes 1–4. The methyl glycoside 3 was synthesized from com-
mercially available 1,2,5,6-diisopropylidene-a-D-glucose in seven
steps according to literature procedures in an overall yield of
9%.18–22 The conversion of the methyl glycoside 3 to the target
compounds 2a–f was achieved over several steps as shown in
Target compound 2g was prepared as shown in Scheme 2. Com-
pound 8,11 was treated with 1,10-thiocarbonyldiimidazole in
Figure 1. Stereochemistry effects comparison of AE (2j), HE (1, 2h and compound 2g) BACE-1 inhibitor Ki values and their caphepsin D selectivity.

C. Björklund et al. / Bioorg. Med. Chem. 18 (2010) 1711–1723
1713
Scheme 1. Reagent and conditions: (i) R1X (12), NaH or Ag2O, DMF, rt; (ii) H2SO4, 1,4-dioxane, reflux; (iii) PDC, DCM, rt; (iv) (S)-2-amino-N-benzyl-3-methyl-butyramide, 2-
hydroxypyridine, DIPEA, 70 °C; (v) Ph3P, H2O, MeOH, rt; (vi) H2, Pd/C, MeOH, rt; (vii) 5-(methanesulfonyl-methyl-amino)-N0-(1-phenyl-ethyl)-isophthalacid, Py-BOP, DIPEA,
DCM, rt.
Scheme 2. Reagents and conditions: (i) TDI, DCE, reflux; (ii) Bu3SnH, AIBN, toluene, reflux; (iii) 5-(methanesulfonyl-methyl-amino)-N0-(1-phenyl-ethyl)-isophthalacid, Py-
BOP, DIPEA, DCM, rt.
Scheme 3. Reagents and conditions: (i) Ph3P, DIAD, para-nitrobenzoic acid, THF, rt; (ii) 0.1 M NaOMe in MeOH; (iii) Ph3P, H2O, MeOH; (iv) 5-(methanesulfonyl-
methylamino)-N0-(1-phenyl-ethyl)-isophthalacid, Py-BOP, DIPEA, DCM, rt.
refluxing dichloroethane, without prior purification the formed
thiocarbonyl ester was subsequently added to tributyltin hydride
and a catalytically amount of AIBN in refluxing toluene, under
these conditions also the azide is reduced as reported by Witczak
and Whistler29 The resulting amine was coupled without prior
purification with 5-(methanesulfonyl-methyl-amino)-N0-(1-phe-
nyl-ethyl)-isophthalacid using Py-BOP and DIPEA in DCM furnish-
ing the target compound 2g in 7% yield over three steps.19,28
The synthesis of target compound 2h is outlined in Scheme 3.
Compound 8 was reacted with Ph3P, diisopropyl azocarboxylate
(DIAD) and 4-nitrobenzoic acid in THF30 furnishing the 4-nitro-
benzoate ester which was hydrolyzed using 0.1 M sodium methox-
ide in methanol delivering the inverted alcohol 9 in 23% yield.
Reduction of the azide group in compound 9 was achieved using
catalytic hydrogenation over palladium on activated carbon in
methanol providing the corresponding amine which was coupled

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C. Björklund et al. / Bioorg. Med. Chem. 18 (2010) 1711–1723
Scheme 4. Reagents and conditions: (i) H2, Pd/C, MeOH, rt; (ii) Boc2O, 1 M NaOH, THF/H2O 1:1; (iii) MsCl, pyridine, 0 °C; (iv) NaN3, DMF, 60 °C; (v) TES, DCM/TFA 2:1, rt; (vi)
5-(methanesulfonyl-methyl-amino)-N0-(1-phenyl-ethyl)-isophthalacid, Py-BOP, DIPEA, DCM, rt; (vii) H2, Pd/C, MeOH, rt.
Figure 2. Diversity reagents 12.
without further purification with 5-(methanesulfonyl-methyl-
amino)-N0-(1-phenyl-ethyl)-isophthalacid using Py-BOP and DIPEA
in DCM furnishing the target compound 2h in 75% yield.
tivity was only a modest 1.6-fold. However, the small size P10 eth-
oxy group of inhibitor 2b retained an excellent fold selectivity of
>1500 over cathepsin D which is comparable with inhibitor 1, hav-
ing a P10 methoxy group. It shows that for this novel HE scaffold
small and polar groups are advantageous for achieving both a high
potency against BACE-1 as well as a high selectivity over cathepsin
D. We have also confirmed the importance of the hydroxyl group
which binds to the catalytic Asp32 and Asp228 for this HE transi-
tion state isostere (see Fig. 1) by preparing both the deoxygenated
analogue 2g and by inverting the configuration of the hydroxyl
group to the R-configuration ((R) CH–OH, 2h), where 2g and 2h dis-
played approximately 1000 and 2000-fold drop in activity, respec-
tively, compared to lead inhibitor 1. These results are consistent
with the hydroxyl group of the HE central core making key hydro-
gen bond interactions to the catalytic Asp228 and Asp32, cf. earlier
reported (R)-NH2 aminoethylene.31 We were also interested in
evaluating the corresponding inhibitor structures where the hy-
droxyl group has been substituted with an amino group furnishing
an aminoethylene (AE) isostere. Inhibitors containing primary
amines have previously been used successfully in two classes of re-
nin inhibitors32,33 and in BACE-1 inhibitors containing either a
aminoethylene (AE) isostere31 or a truncated aminoethylene (AE)
isostere.34 It was found from X-ray crystallography that the pri-
mary amine of these BACE-1 inhibitors was engaging both catalytic
aspartates Asp32 and Asp 228 with hydrogen bonding contacts.
Notably, these AE isostere inhibitor structures delivered compara-
ble BACE-1 inhibitory activities but importantly also bestows the
inhibitors with advantageous physicochemical properties such as
increased solubility and higher affinity for the acidic organelles
wherein BACE-1 and sAPP are co-localized, rendering higher
potencies in the cell-based assays. When the hydroxyl group in
inhibitor 1 was exchanged for an amine, with retention of config-
uration, yielding the corresponding AE inhibitor structure 2j, this
resulted in a BACE-1 Ki value of 140 nM, providing an unexpected
drop in potency in excess of 1000-fold.
Catalytic hydrogenation of 9 over a catalytic amount palladium
on active carbon in methanol followed by subsequent treatment
with di-tert-butyl dicarbonate and sodium hydroxide in a mixture
of THF and H2O gave the Boc-protected compound 10 in 96% yield
(Scheme 4). Compound 10 was converted to the corresponding
azide 11 with inversion of configuration in 25% yield by treating
the alcohol with methane sulfonyl chloride in the presence pyri-
dine followed by displacement of the methane sulfonate group
using sodium azide in DMF at 60 °C for 16 h. Targets compounds
2i and 2j were achieved by deprotecting the Boc group in com-
pound 11 using trifluoroacetic acid and triethyl silane in DCM
followed by coupling of the resulting amine with 5-(methanesulfo-
nyl-methyl-amino)-N0-(1-phenyl-ethyl)-isophthalacid using Py-
BOP and DIPEA in DCM furnishing the target compound 2i in 85%
yield. The corresponding amine 2j was furnished in 85% yield by
catalytic hydrogenation of 2i.
2.2. Structure activity relationships
We have previously reported on the potent lead BACE-1 inhib-
itor 1.11 Based on the encouraging results from inhibitor 1, we have
in a continuation explored a set of P10 groups, all featuring the alk-
oxy motif, to expand on the SAR at this position. These modifica-
tions furnished highly active BACE-1 inhibitors, with Ki values
ranging from 1 to 20 nM (inhibitors 2a–f, Table 1) demonstrating
that the S10-pocket can accommodate larger substituents than pre-
viously anticipated. The S10 pocket in cathepsin D, as in most other
aspartic proteases, comprise of hydrophobic residues, while it is
primarily hydrophilic in BACE-1.14 As expected there was a consid-
erable drop in selectivity over cathepsin D with larger, hydropho-
bic substituents yielding an increase in potencies against
cathepsin D. For the P10 benzyl substituent (inhibitor 2a) the selec-

C. Björklund et al. / Bioorg. Med. Chem. 18 (2010) 1711–1723
1715
Table 1
Target compounds and inhibition data
Entry
Compound
R1
R2
R3
Ki BACE-1b (nM)
Ki Cath Db (nM)
1
2
2a
2b
Bn
Et
OH
OH
H
H
20
2.1
62
3350
O
3
4
5
2c
2d
2e
OH
OH
OH
H
H
H
3.1
3.3
3
240
130
61
Allyl
n-Prop
6
2f
OH
H
1.3
nda
7
8
9
2g
2h
2i
Me
Me
Me
Me
H
H
N3
NH2
H
OH
H
130
210
70
>5000
>5000
>5000
>5000
10
2j
H
140
a
nd = not determined.
Ki values are the mean of at least two different experiments.
b
2.3. Modeling and X-ray crystallography analysis
Other residues in the S10-pocket which are in close contact interac-
tions with the larger P10 groups are Tyr198, Lys224, Ile226, Val332,
and Thr329.
Our earlier published crystal structure of inhibitor 111 shows
two structural water molecules buried in the S10-pocket and a
water mediated hydrogen bond network including the P10-ether
oxygen. The overall binding of compound 2c is the same as for
inhibitor 1 except that at least one of these two water molecules
is displaced by the larger P10 substituents in compounds 2a–f.
The crystal structure of 2c (Fig. 3) shows that the terminal methoxy
oxygen of the P10-tail overlays well with the position of one of the
replaced water molecules. Similarly, as reported earlier for inhibi-
tor 1,11 a structural water molecule forms hydrogen bond linkages
between the inner-oxygen of the P10-residue in 2c and two amino
acid residues in the active site (Fig. 4), namely Thr231 and Arg235.
The variations in the position of the hydroxyl group in the HE
isostere clearly show the importance of having a hydrogen bond
network with both Asp32 and Asp228. The hydroxyl group of
inhibitor 1 and 2a–f is perfectly positioned in between the aspartic
acid side chains, whereas modeling of the inverted alcohol 2h
shows interactions with only one of the aspartic acids (Asp32).
The low activity of the amine 2j cannot be readily explained from
modeling and could be due to a shift in the degree of protonation of
the amine in the active site, due to electronegative effects from the
P1 aryloxy and P10 methoxy substituents, which would render less
favorable binding to the catalytic Asp32 and Asp228. It could also
Figure 3. X-ray structure of inhibitor 2c showing the flexibility of the P10-tail and the hydrogen bond network with a structural water molecule including the inner-ether
oxygen in the ligand and the residues Thr231 and Arg235.

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otherwise, all materials were obtained from commercial suppliers
and used without further purification. DCM was refluxed over
CaH2 and distilled onto 4 Å MS before use. Compounds 2a–j were
further purified before the biological testing using preparative RP-
HPLC, consisted of Waters 2767 auto-injector and fraction collector,
Waters996 photodiodearray detector, and MicromassZQ2000 mass
detector (operated in +ESI). The preparative reversed phase column
was an ACE C8, 21 ꢁ 100 mm, 5
lm, 100A from ACE (UK) and the mo-
bile phases were based on water/acetonitrile containing 0.1%TFA.
Optical rotations were measured at room temperature on a Per-
kin–Elmer 341 polarimeter using a 10 cm, 1 mL cell.
4.5. LC–MS purity measurements
Figure 4. Overview of the hydrogen bonding network of inhibitor 2c.
4.5.1. Chromatography system A
Column: ACE C8, 50 ꢁ 3 mm, 3
lm particles; pump: Waters Alli-
be attributed to a delicate balance between the over-all fit of these
relatively large inhibitor structures and fine-tuned geometrical
constraints in the active site making these inhibitors sensitive to
small structural variations around the active site.
ance 2695; UV/vis-detector: Waters 996; MS detector: Waters ZQ;
mobil phase A: 10 mM NH4OAc in water; Mobil phase B: 10 mM
NH4OAc in 90% acetonitrile; gradient: 20–100% B in 5 min followed
by 2 min at 100% B.
3. Conclusions
4.5.2. Chromatography system B
As chromatography system A except: mobil phase A: 0.2%
HCOOH in water; mobil phase B: 0.18% HCOOH in 100%
acetonitrile.
This study demonstrates that both potent and selective BACE-1
inhibitors can be achieved by exploiting the unique features of the
S10 pocket in BACE-1 with the newly developed HE template. Fur-
ther investigation of this inhibitor series by reducing molecular
weight and retaining the favorable structural elements is highly
warranted and these efforts on inhibitor design and with cell-
based activity data are forthcoming.
4.6. Synthetic procedures
4.6.1. Methyl 5-azido-3,5-dideoxy-6-O-(3,5-difluoryphenyl)-
L-
lyxo-hexofuranoside (3)
The methyl glycoside 3 was synthesized from commercially
available 1,2,5,6-diisopropylidene-a-D-glucose in seven steps
4. Experimental section
according to literature procedures in an overall yield of 9%.18–22
1H NMR (400 MHz, CDCl3) d 6.49–6.41 (m, 3H), 4.85 (s, 1H),
4.56–4.50 (m, 1H), 4.30 (t, JHH = 4.5 Hz, 1H), 4.10–4.01 (m, 2H),
3.73–3.67 (m, 1H), 3.41 (s, 3H), 2.17–2.09 (m, 1H), 2.02–1.95 (m,
1H), 1.72 (d, JHH = 4.6 Hz, 1H); 13C NMR (100.5 MHz, CDCl3) d
4.1. Protease enzyme assay
The BACE1 and cathepsin D assays were performed as previ-
ously described.35
1
1
163.9 (d, JCF = 247.5 Hz), 163.7 (d, JCF = 247.5 Hz), 160.1 (t,
2
4.2. Crystallography
3JCF = 13.7 Hz), 109.6, 98.6 (d, JCF = 28.9 Hz, 2C), 97.2 (t,
2JCF = 25.9 Hz), 79.0, 75.8, 68.6, 64.7, 55.2, 35.0; MS (ESI) m/z
þ
The details of the crystallization and structure determination
procedures have been published elsewhere.35 Briefly, the complex
of BACE-1 and 2c was crystallized in the monoclinic space group
P21 with cell dimensions of a = 81.4, b = 102.5, c = 100.2 and
b = 103.5°. The structure was determined to 2.1 Å resolution with
an R-value of 0.21 (R-free 0.24) and deposited in the RCSB PDB data
base (3125).
338.1 [(M+Na)+ calcd for C13H15F2N3NaO4 338.1].
4.6.2. Methyl 5-azido-2-O-benzyl-3,5-dideoxy-6-O-(3,5-
difluorophenyl)-L-lyxo-hexofuranoside (4a)
Compound 3 (226 mg, 0.717 mmol) was dissolved in DMF
(5 mL), and benzyl bromide (0.68 mL, 5.76 mmol, 8 equiv) and
Ag2O (332 mg, 1.43 mmol, 2 equiv) were added. The reaction was
stirred at room temperature over night. The reaction was quenched
with CHCl3 and the solids were filtered off. The filtrate was concen-
trated under vacuum and the residue was purified by column chro-
matography (isohexane?isohexane/ethyl acetate 10:1) to yield
compound 4a (136 mg, 0.337 mmol, 47%). 1H NMR (400 MHz,
CDCl3) d 7.40–7.27 (m, 5H), 6.50–6.41 (m, 3H), 4.98 (s, 1H), 4.53
(d, JHH = 5.8 Hz, 2H), 4.55–4.48 (m, 1H), 4.10–4.01 (m, 3H), 3.73–
4.3. Modeling
All modeling experiments were performed using SYBYL 8.0 (Tri-
pos Inc. 1699 South Hanley Road, St. Louis, Missouri, 63144, USA).
4.4. General methods
3.67 (m, 1H), 3.41 (s, 3H), 2.11–2.06 (m, 2H); 13C NMR
1
All glassware were dried over an open flame before use in con-
nection with an inert atmosphere. Concentrations were performed
under reduced pressure at <40 °C (bath temperature). Thin layer
chromatography was performed using Merck Silica Gel 60 F-254
plates with detection by UV, charring with 8% sulfuric acid or
ammonium molybdate (100 g): Ce(IV)sulfate (2 g): sulfuric acid
(10%, 2 L). Column chromatography was performed on silica
(0.035–0.070 mm). NMR spectra were recorded at 25 °C on a Var-
ian (400 MHz) or on a Bruker (400 MHz or 500 MHz) instrument
using the solvent residual peak (CDCl3 1H d 7.26 and 13C d 77.17
or CD3OD-d4 1H d 3.31 and 13C 49.0) as standard. Unless stated
(100.5 MHz, CDCl3)
d
163.9 (d, JCF = 246.7 Hz), 163.7 (d,
1JCF = 246.7 Hz), 160.2 (t, JCF = 13.3 Hz), 137.7, 128.7, 128.0,
3
2
2
127.8, 107.3, 98.6 (d, JCF = 28.7 Hz, 2C), 97.2 (t, JCF = 25.8 Hz),
82.8, 79.4, 71.6, 68.7, 64.4, 55.2, 32.8; MS (ESI) m/z 428.0
([M+Na]+ calcd for C20H21F2N3NaO4 428.1).
þ
4.6.3. Methyl 5-azido-3,5-dideoxy-6-O-(3,5-difluorophenyl)-2-
O-ethyl-L-lyxo-hexofuranoside (4b)
Compound 3 (248 mg, 0.787 mmol) was dissolved in DMF
(5 mL), and ethyl iodide (0.50 mL, 6.29 mmol, 8 equiv) and Ag2O
(365 mg, 1.57 mmol, 2 equiv) were added. The reaction was stirred

C. Björklund et al. / Bioorg. Med. Chem. 18 (2010) 1711–1723
1717
at room temperature over night. The reaction mixture was
quenched with CHCl3 and the solids were filtered off. The filtrate
was concentrated under vacuum and the residue was purified by
column chromatography (isohexane?isohexane/ethyl acetate
20:1) to yield compound 4b (92 mg, 0.267 mmol, 34%). 1H NMR
(400 MHz, CDCl3) d 6.49–6.40 (m, 3H), 4.91 (s, 1H), 4.50–4.43 (m,
1H), 4.10–4.01 (m, 2H), 3.90 (t, JHH = 2.8 Hz, 1H), 3.72–3.66 (m,
1H), 3.58–3.49 (m, 2H), 3.40 (s, 3H), 2.06–2.01 (m, 2H), 1.21 (t,
3 equiv) dissolved in DMF (4 mL) was added drop wise. The reac-
tion was stirred in room temperature over night and then
quenched with methanol, concentrated and purified by column
chromatography (toluene/ethyl acetate 10:1) to yield compound
4e (164 mg, 0.44 mmol, 23%). 1H NMR (400 MHz, CDCl3) d 6.48–
6.40 (m, 3H), 4.92 (s, 1H), 4.52–4.40 (m, 1H), 4.10–4.00 (m, 2H),
3.74–3.65 (m, 1H), 3.40 (s, 3H), 3.37–3.35 (m, 2H), 3.34–3.30 (m,
1H), 2.07–2.01 (m, 2H), 0.57–0.51 (m, 2H), 0.23–0.17 (m, 2H);
1
JHH = 7.0 Hz, 3H); 13C NMR (100.5 MHz, CDCl3)
d
163.9 (d,
13C NMR (100.5 MHz, CDCl3) d 163.9 (d, JCF = 246.8 Hz), 163.7 (d,
1
3
1JCF = 246.7 Hz), 163.7 (d, JCF = 246.7 Hz), 160.2 (t, JCF = 13.3 Hz),
1JCF = 246.8 Hz), 160.2 (t, 3JCF = 13.7 Hz), 106.8, 98.6 (d,
2
2
2
107.3, 98.4 (d, JCF = 28.7 Hz, 2C), 97.2 (t, JCF = 25.8 Hz), 83.0,
79.3, 68.8, 65.1, 64.5, 55.3, 32.8, 15.5; MS (ESI) m/z 366.0
2JCF = 28.5 Hz, 2C), 97.2 (t, JCF = 25.9 Hz), 84.7, 79.2, 68.7, 64.4,
57.1, 55.2, 32.3, 10.8, 3.3, 3.2; MS (ESI) m/z 392.3 ([M+Na]+ calcd
([M+Na]+ calcd for C15H19F2N3NaO4 366.1).
for C17H21F2N3NaO4 392.1).
þ
þ
4.6.4. Methyl 5-azido-3,5-dideoxy-6-O-(3,5-difluorophenyl)-2-
4.6.7. 5-Azido-2-O-benzyl-3,5-dideoxy-6-O-(3,5-
O-(2-methoxy-ethoxy)-L-lyxo-hexofuranoside (4c)
difluorophenyl)-L-lyxo-hexofuranose (5a)
Compound 3 (161 mg, 0.511 mmol) dissolved in DMF (1 mL)
was added to 60% sodium hydride (31 mg, 0.766 mmol, 1.5 equiv)
dissolved in DMF (1 mL) at 0 °C. The reaction mixture was stirred
for 30 min before 2-bromoethyl methyl ether (0.38 mL, 4.09 mmol,
8 equiv) was added at 0 °C. The reaction was then stirred in room
temperature for 3 h and then quenched with methanol, concen-
trated and purified by column chromatography (isohexane?iso-
hexane/ethyl acetate 20:1) to yield compound 4c (37 mg,
0.099 mmol, 19%). 1H NMR (400 MHz, CDCl3) d 6.50–6.39 (m,
3H), 4.94 (s, 1H), 4.52–4.42 (m, 1H), 4.08–4.01 (m, 2H), 3.97–3.94
(m, 1H), 3.71–3.66 (m, 1H), 3.66–3.62 (m, 2H), 3.40 (s, 3H), 3.38
(s, 3H), 2.13–2.00 (m, 2H); 13C NMR (100.5 MHz, CDCl3) d 163.8
Compound 4a (136 mg, 0.336 mmol) was dissolved in 1,4-diox-
ane:0.5 M H2SO4 1:1 (14 mL) and heated to reflux. After complete
reaction (ꢀ1 h according to TLC), the reaction was cooled to room
temperature and thenneutralizedwith Na2CO3 (aq). Thevolatile sol-
vents were concentrated under vacuum. The residue was dissolved
in DCM and washed with H2O (ꢁ2). The organic phasewas driedover
Na2SO4 and concentrated. The residue was purified by column chro-
matography (isohexane?isohexane/ethyl acetate 10:1) to yield
compound 5a (61 mg, 0.157 mmol, 47%). 1H NMR (400 MHz, CDCl3)
d7.39–7.27 (m, 5H), 6.50–6.41 (m, 3H), 5.39 (d, JHH = 6. Hz, 1H), 4.58
(d, JHH = 7.0 Hz, 2H), 4.55–4.49 (m, 1H), 4.20–4.16 (m, 2H), 4.04–4.01
(m, 1H), 3.83–3.78 (m, 1H), 3.03–3.00 (m, 1H), 2.25–2.09 (m, 2H);
1
1
3
1
(d, JCF = 246.7 Hz), 163.7 (d, JCF = 246.7 Hz), 160.2 (t, JCF
13.3 Hz), 107.2, 98.6 (d, JCF = 28.7 Hz, 2C), 97.2 (t, JCF = 25.8 Hz),
83.6, 79.3, 72.0, 69.0, 68.7, 64.4, 59.3, 55.3; MS (ESI) m/z 396.0
=
13C NMR (100.5 MHz, CDCl3) d 163.9 (d, JCF = 246.8 Hz), 163.7 (d,
2
2
1JCF = 246.8 Hz), 160.0 (t, 3JCF = 13.3 Hz), 137.7, 128.7, 128.1, 127.8,
100.9, 98.6 (d, 2JCF = 28.7 Hz, 2C), 97.3 (t, 2JCF = 25.7 Hz), 83.6, 78.8,
71.6, 69.6, 63.4, 32.1; MS (ESI) m/z 414.0 ([M+Na]+ calcd for
([M+Na]+ calcd for C16H21F2N3NaO5 396.1).
þ
þ
C19H19F2N3NaO4 414.1).
4.6.5. Methyl 2-O-allyl-5-azido-3,5-dideoxy-6-O-(3,5-
difluorophenyl)-
L
-lyxo-hexofuranoside (4d)
4.6.8. 5-Azido-3,5-dideoxy-6-O-(3,5-difluorophenyl)-2-O-ethyl-
Compound 3 (279 mg, 0.884 mmol) dissolved in DMF (2 mL)
was added to 60% sodium hydride (31 mg, 0.766 mmol, 1.5 equiv)
dissolved in DMF (4 mL) at 0 °C. The reaction mixture was stirred
for 20 min before allyl bromide (0.61 mL, 7.07 mmol, 8 equiv)
was added drop wise at 0 °C. The reaction was stirred in room tem-
perature over night and then quenched with methanol, concen-
trated and purified by column chromatography (isohexane?
isohexane/ethyl acetate 20:1) to yield compound 4d (155 mg,
0.435 mmol, 49%). 1H NMR (400 MHz, CDCl3) d 6.49–6.40 (m,
3H), 5.95–5.82 (m, 1H), 5.33–5,24 (m, 1H), 5.23–5.17 (m, 1H),
4.93 (s, 1H), 4.51–4.43 (m, 1H), 4.08–4.04 (m, 2H), 4.04–4.01 (m,
2H), 3.96 (t, JHH = 2.8 Hz, 1H), 3.72–3.66 (m, 1H), 3.40 (s, 3H),
2.05 (dd, JHH = 7.8, 2.8 Hz, 2H); 13C NMR (100.5 MHz, CDCl3) d
L
-lyxo-hexofuranose (5b)
Compound 5b (72 mg, 0.219 mmol, 82%) was synthesized from
4b (92 mg, 0.267 mmol) according to the method for the prepara-
tion of 5a. 1H NMR (400 MHz, CDCl3) d 6.50–6.39 (m, 3H), 5.32 (d,
JHH = 4.4 Hz, 1H), 4.51–4.44 (m, 1H), 4.19–4.14 (m, 2H), 3.90 (d,
JHH = 4.7 Hz, 1H), 3.82–3.77 (m, 1H), 3.57–3.52 (m, 2H), 3.07 (d,
JHH = 5.4 Hz, 1H), 2.22–2.05 (m, 2H), 1.20 (t, JHH = 7.0 Hz, 3H); 13C
1
NMR (100.5 MHz, CDCl3) d 164.0 (d, JCF = 246.8 Hz), 162.8 (d,
1JCF = 246.8 Hz), 160.0 (t, 3JCF = 13.3 Hz), 100.9, 98.6 (d,
2JCF = 28.7 Hz, 2C), 97.3 (t, JCF = 25.7 Hz), 83.8, 78.7, 69.5, 65.1,
2
63.5, 32.1, 15.5; MS (ESI) m/z 352.0 ([M+Na]+ calcd for
þ
C14H17F2N3NaO4 352.1).
1
1
163.9 (d, JCF = 246.8 Hz), 163.7 (d, JCF = 246.8 Hz), 160.0 (t,
4.6.9. 5-Azido-3,5-dideoxy-6-O-(3,5-difluorophenyl)-2-O-(2-
methoxy-ethoxy)-L-lyxo-hexofuranose (5c)
2
3JCF = 13.3 Hz), 134.1, 117.4, 107.2, 98.4 (d, JCF = 28.7 Hz, 2C),
2
97.1 (t, JCF = 25.7 Hz), 82.5, 79.2, 70.4, 68.6, 64.3, 55.1, 32.6; MS
Compound 5c (49 mg, 0.136 mmol, 80%) was synthesized from
4c (64 mg, 0.170 mmol) according to the method for the prepara-
tion of 5a. 1H NMR (400 MHz, CDCl3) d 6.49–6.39 (m, 3H), 5.35
(d, JHH = 5.8 Hz, 1H), 4.52–4.43 (m, 1H), 4.15 (d, JHH = 6.0 Hz, 2H),
3.96–3.93 (m, 1H), 3.82–3.75 (m, 1H), 3.67–3.62 (m, 2H), 3.54–
3.50 (m, 2H), 3.39 (s, 1H), 3.37 (s, 3H), 3.26 (d, JHH = 5.8 Hz, 1H)
2.22–2.08 (m, 2H); 13C NMR (100.5 MHz, CDCl3) d 163.9 (d,
(ESI) m/z 378.1 ([M+Na]+ calcd for C16H19F2N3NaO4 378.1).
þ
4.6.6. Methyl 5-azido-2-O-cyclopropylmethyl-3,5-dideoxy-6-O-
(3,5-difluorophenyl)-L-lyxo-hexofuranoside (4e)
Hydroxymethyl-cyclopropane (5.92 mmol, 0.47 mL) was dis-
solved in pyridine (5 mL) and treated with tosylchloride (2.26 g,
11.84 mmol, 2 equiv) at 0 °C. After 30 min the reaction was
quenched with methanol, diluted with DCM and washed with
10% NaCl (aq) (2ꢁ 20 mL). The organic phase was dried over
Na2SO4, filtered and concentrated under vacuum. The tosylated
alcohol was then used in the alkylating step without further puri-
fication. Compound 3 (622 mg, 1.97 mmol) dissolved in DMF
(4 mL) was added to 60% sodium hydride (118 mg, 2.96 mmol,
1.5 equiv) dissolved in DMF (2 mL) at 0 °C. The reaction mixture
was stirred for 30 min before the tosylated alcohol (5.92 mmol,
1JCF = 246.8 Hz), 163.7 (d, JCF = 246.8 Hz), 160.0 (t, JCF = 13.3 Hz),
1
3
2
2
100.8, 98.6 (d, JCF = 28.6 Hz, 2C), 97.3 (t, JCF = 25.7 Hz), 84.5,
78.7, 72.0, 69.5, 69.0, 63.5, 59.2, 32.0; MS (ESI) m/z 382.0
([M+Na]+ calcd for C15H19F2N3NaO5 382.1).
þ
4.6.10. 2-O-Allyl-5-azido-3,5-dideoxy-6-O-(3,5-difluorophenyl)-
L-lyxo-hexofuranose (5d)
Compound 5d (82 mg, 0.241 mmol, 55%) was synthesized from
4d (155 mg, 0.435 mmol) according to the method for the prepara-

1718
C. Björklund et al. / Bioorg. Med. Chem. 18 (2010) 1711–1723
2
tion of 5a. 1H NMR (400 MHz, CDCl3) d 6.51–6.39 (m, 3H), 5.97–
5.81 (m, 1H), 5.36–5.32 (m, 1H), 5.32–5.25 (m, 1H), 5.22–5.17
(m, 1H), 4.53–4.45 (m, 1H), 4.17 (d, JHH = 6.2 Hz, 2H), 4.06–4.02
(m, 2H), 3.96 (d, JHH = 4.8 Hz, 1H), 3.15 (d, JHH = 5.8 Hz, 1H), 2.24–
2C), 97.6 (t, JCF = 25.7 Hz), 75.8, 73.8, 71.8, 70.1, 68.6, 62.8,
59.2, 32.6; MS (ESI) m/z 358.0 ([M+H]+ calcd for C15H18F2N3O5
þ
358.1).
2.04 (m, 2H); 13C NMR (100.5 MHz, CDCl3)
d
163.9 (d,
4.6.15. 2-O-Allyl-5-azido-3,5-dideoxy-6-O-(3,5-difluorophenyl)-
-lyxo-1,4-lactone (6d)
1
3
1JCF = 246.8 Hz), 163.7 (d, JCF = 246.8), 160.0 (t, JCF = 13.3 Hz),
L
2
134.2, 117.6, 100.9, 98.6 (d, JCF = 28.7 Hz, 2C), 97.3 (t,
Compound 6d (31 mg, 0.092 mmol, 38%) was synthesized from
2JCF = 25.7 Hz), 83.4, 78.7, 70.5, 69.6, 63.5, 32.0; MS (ESI) m/z
5d (82 mg, 0.241 mmol) according to the method for the prepara-
tion of 6a. 1H NMR (400 MHz, CDCl3) d 6.52–6.41 (m, 3H), 5.97–
5.85 (m, 1H), 5.38–5.24 (m, 2H), 4.82–4.77 (m, 1H), 4.44–4.37
(m, 1H), 4.34 (dd, JHH = 5.8, 8.2 Hz 1H), 4.23 (d, JHH = 6.3 Hz, 2H),
4.22–4.16 (m, 1H), 3.94–3.89 (m, 1H), 2.57–2.48 (m, 1H), 2.44–
2.35 (m, 1H); 13C NMR (100.5 MHz, CDCl3) d 173.9, 163.9 (d,
364.1 ([M+Na]+ calcd for C15H17F2N3NaO4 364.1).
þ
4.6.11. 5-Azido-2-O-cyclopropylmethyl -3,5-dideoxy-6-O-(3,5-
difluorophenyl)-L-lyxo-hexofuranose (5e)
Compound 5e (47 mg, 0.132 mmol, 67%) was synthesized from
4e (73 mg, 0.198 mmol) according to the method for the prepara-
tion of 5a. 1H NMR (400 MHz, CDCl3) d 6.49–6.41 (m, 3H), 5.33
(br s, 1H), 4.53–4.47 (m, 1H), 4.19–4.16 (m, 2H), 3.95–3.92 (m,
1H), 3.82–3.77 (m, 1H), 3.38–3.31 (m, 2H), 2.24–2.05 (m, 2H),
1.30–1.22 (m, 1H), 1.14–0.98 (m, 1H), 0.63–0.51 (m, 2H), 0.26–
1JCF = 247.1 Hz), 162.7 (d, JCF = 247.1 Hz), 159.7 (t, JCF = 13.7 Hz),
1
3
2
2
133.6, 118.6, 98.7 (d, JCF = 28.4 Hz, 2C), 97.7 (t, JCF = 25.8 Hz),
75.8, 72.4, 71.6, 68.6, 62.8, 32.7; MS (ESI) m/z 362.1 ([M+Na]+ calcd
þ
for C15H15F2N3NaO4 362.1).
0.18 (m, 2H); 13C NMR (100.5 MHz, CDCl3)
d
163.9 (d,
4.6.16. 5-Azido-2-O-cyclopropylmethyl-3,5-dideoxy-6-O-(3,5-
difluorophenyl)-L-lyxo-1,4-lactone (6e)
1
3
1JCF = 246.7 Hz), 163.7 (d, JCF = 246.7 Hz), 160.0 (t, JCF = 13.6 Hz),
2
2
101.0, 98.6 (d, JCF = 28.4 Hz, 2C), 97.3 (t, JCF = 25.8 Hz), 83.7,
78.7, 74.5, 69.6, 63.5, 32.1, 10.8, 3.4, 3.3; MS (ESI) m/z 378.1
Compound 6e (39 mg, 0.110 mmol, 83%) was synthesized from
5e (47 mg, 0.132 mmol) according to the method for the prepara-
tion of 6a. 1H NMR (400 MHz, CDCl3) d 6.53–6.41 (m, 3H), 4.82–
4.77 (m, 1H), 4.33 (dd, JHH = 5.8, 8.2 Hz, 1H), 4.22 (d, JHH = 6.3 Hz,
2H), 3.95–3.89 (m, 1H), 3.71 (dd, JHH = 7.2, 10.0 Hz, 1H), 3.49 (dd,
JHH = 6.9, 10.2 Hz, 1H), 2.57–2.49 (m, 1H), 2.45–2.36 (m, 1H),
1.14–1.03 (m, 1H), 0.62–0.51 (m, 2H), 0.30–0.21 (m, 2H); 13C
([M+Na]+ calcd for C16H19F2N3NaO4 378.1).
þ
4.6.12. 5-Azido-2-O-benzyl-3,5-dideoxy-6-O-(3,5-
difluorophenyl)-L-lyxo-1,4-lactone (6a)
Compound 5a (61 mg, 0.157 mmol) was dissolved in DCM
(5 mL). At 0 °C pyridinium dichromate (88 mg, 0.235 mmol,
1.5 equiv) and 4 Å molecular sieves powder were added. The reac-
tion was stirred over night in room temperature. The solids were
filtered off and the filtrate was concentrated under vacuum, the
residue was purified by column chromatography (isohexane?iso-
hexane/ethyl acetate 15:1) to yield compound 6a (59 mg,
0.150 mmol, 96%). 1H NMR (400 MHz, CDCl3) d 7.41–7.30 (m,
5H), 6.52–6.40 (m, 3H), 4.96 (d, 1H, JHH = 11.6 Hz), 4.82–4-76 (m,
1H), 4.70 (d, 1H, JHH = 11.6 Hz), 4.36 (dd, JHH = 5.7, 8.3 Hz, 1H),
4.22 (d, JHH = 6.3 Hz, 2H), 3.92–3.87 (m, 1H), 2.53–2.35 (m, 2H);
1
NMR (100.5 MHz, CDCl3) d 174.0, 163.9 (d, JCF = 247.2 Hz), 163.7
(d, JCF = 247.2 Hz), 159.7 (t, JCF = 13.7 Hz), 98.6 (d, JCF = 28.8 Hz,
1
3
2
2
2C), 97.7 (t, JCF = 25.8 Hz), 75.9, 75.7, 73.0, 68.6, 62.8, 32.8, 10.6,
3.4, 3.1; MS (ESI) m/z 376.1 ([M+Na]+ calcd for C16H17F2N3NaO4
þ
376.1).
4.6.17. 5-Azido-2-benzyloxy-6-(3,5-difluoro-phenoxy)-4-
hydroxy-hexanoid acid (1-benzylcarbamoyl-2-methyl-propyl)-
amide (7a)
Compound 6a (43 mg, 0.111 mmol) and (S)-2-amino-N-benzyl-
3-methyl-butyramide (69 mg, 0.333 mmol, 3 equiv) were dis-
solved in diisopropyl ethyl amine (5 mL). 2-Hydroxy-pyridine
(21 mg, 0.222, 2 equiv) was added and the mixture was heated to
70 °C over night, if needed a few drops of DMF was added for sol-
ubility. The reaction mixture was concentrated and the residue
was purified by column chromatography (isohexane/ethyl acetate
10:1) to give compound 7a (46 mg, 0.078 mmol, 70%). 1H NMR
(400 MHz, CDCl3) d 7.41–7.20 (m, 10H), 7.10 (d, JHH = 8.8 Hz, 1H),
6.87 (t, JHH = 5.6 Hz, 1H), 6.50–6.35 (m, 3H), 4.64 (d, JHH = 11.6 Hz,
1H), 4.59 (d, JHH = 11.6 Hz, 1H) 4.41 (d, JHH = 5.6 Hz, 2H), 4.31 (dd,
JHH = 5.2, 8.8 Hz, 1H), 4.14–4.08 (m, 1H), 4.03–3.98 (dd, JHH = 4.2,
9.8 Hz, 1H), 3.97–3.90 (m, 2H), 3.64–3.55 (m, 1H), 2.55 (d,
JHH = 7.6 Hz, 1H), 2.47–2.35 (m, 1H), 2.25–2.13 (m, 1H), 1.94–1.85
(m, 1H), 0.96 (d, JHH = 6.9 Hz, 3H), 0.91 (d, JHH = 6.9 Hz, 3H); 13C
1
13C NMR (100.5 MHz, CDCl3) d 173.8, 163.9 (d, JCF = 247.3 Hz)
1
3
163.7 (d, JCF = 247.3 Hz), 159.5 (t, JCF = 13.3 Hz), 136.8, 128.7,
2
2
128.2, 98.5 (d, JCF = 28.7 Hz, 2C), 97.5 (t, JCF = 25.7 Hz), 75.9,
72.6, 72.4, 68.5, 62.7, 32.7; MS (ESI) m/z 412.0 ([M+Na]+ calcd for
þ
C19H17F2N3NaO4 412.1).
4.6.13. 5-Azido-3,5-dideoxy-6-O-(3,5-difluorophenyl)-2-O-
ethyl-L-lyxo-1,4-lactone (6b)
Compound 6b (49 mg, 0.150 mmol, 68%) was synthesized from
5b (72 mg, 0.219 mmol) according to the method for the prepara-
tion of 6a. 1H NMR (400 MHz, CDCl3) d 6.50–6.41 (m, 3H), 4.81–
4.73 (m, 1H), 4.27 (dd, JHH = 5.8, 8.2 Hz, 1H), 4.22 (d, JHH = 6.4 Hz,
2H), 3.98–3.88 (m, 2H), 3.68–3.59 (m, 1H), 2.55–2.47 (m, 1H),
2.40–2.30 (m, 1H), 1.23 (t, JHH = 7.0 Hz, 3H); 13C NMR
1
1
(100.5 MHz, CDCl3) d 174.0, 163.9 (d, JCF = 247.3 Hz), 163.7 (d,
NMR (100.5 MHz, CDCl3) d 172.6, 170.8, 163.9 (d, JCF = 246.8 Hz),
163.7 (d, JCF = 246.8 Hz), 160.1 (t, JCF = 13.6 Hz), 138.2, 136.6,
129.0, 128.9, 128.8, 128.1, 128.0, 127.7, 98.7 (d, JCF = 28.7 Hz,
3
2
1
3
1JCF = 247.3 Hz), 159.7 (t, JCF = 13.3 Hz), 98.6 (d, JCF = 28.7 Hz,
2
2
2C), 97.6 (t, JCF = 25.7 Hz), 75.9, 73.3, 68.6, 66.6, 62.8, 32.7, 15.2;
MS (ESI) m/z nd ([M+Na]+ calcd for C14H15F2N3NaO4 350.1).
2C), 97.3 (t, JCF = 25.8 Hz), 77.0, 73.1, 68.9, 67.3, 64.7, 58.3, 43.8,
2
þ
36.0, 29.9, 19.8, 17.6; MS (ESI) m/z 618.1 ([M+Na]+ calcd for
þ
4.6.14. 5-Azido-3,5-dideoxy-6-O-(3,5-difluorophenyl)-2-O-(2-
C31H35F2N5NaO5 618.3).
methoxy-ethoxy)-L-lyxo-1,4-lactone (6c)
Compound 6c (40 mg, 0.111 mmol, 81%) was synthesized from
5c (49 mg, 0.136 mmol) according to the method for the prepara-
tion of 6a. 1H NMR (400 MHz, CDCl3) d 6.51–6.40 (m, 3H), 4.81–
4.74 (m, 1H), 4.35 (dd, JHH = 6.0, 8.5 Hz, 1H), 4.21 (d, JHH = 6.4 Hz,
2H), 4.12–4.06 (m, 1H), 3.94–3.88 (m, 1H), 3.81–3.74 (m, 1H),
3.59–3.54 (m, 1H), 3.38 (s, 3H), 2.58–2.39 (m, 2H); 13C NMR
4.6.18. 5-Azido-6-(3,5-difluoro-phenoxy)-2-etoxy-4-hydroxy-
hexanoid acid (1-benzylcarbamoyl-2-methyl-propyl)-amide
(7b)
Compound 7b (67 mg, 0.126 mmol, 84%) was synthesized from
6b (49 mg, 0.150 mmol) according to the method for the prepara-
tion of 7a. 1H NMR (400 MHz, CDCl3) d 7.32–7.18 (m, 5H), 7.17–
7.06 (m, 2H), 6.47–6.35 (m, 3H), 4.40 (t, JHH = 5.1 Hz, 2H), 4.31
(dd, JHH = 5.4, 9.1 Hz, 1H), 4.05 (dd, JHH = 4.0, 9.6 Hz, 1H), 3.98–
1
(100.5 MHz, CDCl3) d 173.9, 163.9 (d, JCF = 247.3 Hz), 163.7 (d,
3
2
1JCF = 247.3 Hz), 159.7 (t, JCF = 14.0 Hz), 98.6 (d, JCF = 29.5 Hz,

C. Björklund et al. / Bioorg. Med. Chem. 18 (2010) 1711–1723
1719
3.90 (m, 3H), 3.66–3.52 (m, 3H), 3.04 (br s, 1H), 2.41–2.30 (m,
1H), 2.15–2.04 (m, 1H), 1.91–1.83 (m, 1H), 1.23 (t, JHH = 7.0 Hz,
3H), 0.96 (d, JHH = 6.8 Hz, 3H), 0.92 (d, JHH = 6.8 Hz, 3H); 13C
4.6.22. N-[(1S,2S,4R)-4-((S)-1-Benzylcarbamoyl-2-methyl-
propylcarbamoyl)-4-benzyl-1-(3,5-difluoro-phenoxymethyl)-2-
hydroxy-butyl]-5-(methanesulfonyl-methyl-amino)-N0-((R)-1-
phenyl-ethyl)-isophthalamide (2a)
Azide 7a (46 mg, 0.078 mmol) and triphenyl phosphine (31 mg,
0.012 mmol, 1.5 equiv) were dissolved in MeOH (5 mL) and four
drops of water were added. The reaction was stirred at room temper-
ature over night and then concentrated under vacuum. Without fur-
ther purification the formed amine was used in the next step. 5-
(Methanesulfonyl-methyl-amino)-N0-(1-phenyl-ethyl)-isophthal-
acid (29 mg, 0.078 mmol, 1 equiv), Py-BOP (40 mg, 0.078 mmol,
1
NMR (100.5 MHz, CDCl3) d 172.8, 171.0, 163.8 (d, JCF = 246.7 Hz),
1
3
163.7 (d, JCF = 246.7 Hz), 160.1 (t, JCF = 13.7 Hz), 138.3, 128.8,
2
2
127.9, 127.6, 98.6 (d, JCF = 29.1 Hz, 2C), 97.1 (t, JCF = 25.7 Hz),
77.9, 68.9, 67.6, 66.8, 64.7, 58.1, 43.6, 36.2, 30.1, 19.7, 17.6,
15.5; MS (ESI) m/z 556.1 ([M+Na]+ calcd for C26H33F2N5NaO5
þ
556.2).
4.6.19. 5-Azido-6-(3,5-difluoro-phenoxy)-4-hydroxy-2-(2-
methoxy-etoxy)-hexanoid acid (1-benzylcarbamoyl-2-methyl-
propyl)-amide (7c)
1 equiv) and DIPEA (14 lL, 0.078 mmol, 1 equiv) were dissolved in
DCM (2.5 mL). The mixture was stirred at room temperature for
Compound 7c (54 mg, 0.096 mmol, 87%) was synthesized from
6c (40 mg, 0.110 mmol) according to the method for the prepara-
tion of 7a.
30 min before the amine (ꢀ1 equiv) from the previous reaction dis-
solved in DCM (4 mL) and DIPEA (14 lL, 0.078 mmol, 1 equiv) were
added. After complete reaction the mixture were washed with NaH-
CO3 (1ꢁ 10 mL) and brine (1ꢁ 10 mL). The water phase was ex-
tracted with DCM (2ꢁ 10 mL). The organic layers were combined
and dried over Na2SO4, concentrated under vacuum and purified
by column chromatography (isohexane/ethyl acetate 1:1) to give
1H NMR (400 MHz, CDCl3) d 7.45 (d, JHH = 8.8 Hz, 1H), 7.35–7.20
(m, 5H), 7.00 (t, JHH = 5.6 Hz, 1H), 6.49–6.37 (m, 3H), 4.41 (dd,
JHH = 3.5, 5.8 Hz, 2H), 4.29 (dd, JHH = 5.9, 8.6 Hz, 1H), 4.06 (dd,
JHH = 4.0, 9.7 Hz, 1H), 4.01–3.93 (m, 3H), 3.76–3.61 (m, 3H), 3.59–
3.48 (m, 2H), 3.38 (s, 3H), 3.05 (d, JHH = 6.8 Hz, 1H), 2.41–2.29
(m, 1H), 2.17–2.06 (m, 1H), 1.96–1.87 (m, 1H), 0.98 (d, JHH = 6.8 Hz,
3H), 0.95 (d, JHH = 6.8 Hz, 3H); 13C NMR (100.5 MHz, CDCl3) d 172.7,
171.0, 163.8 (d, 1JCF = 246.6 Hz), 163.7 (d, 1JCF = 246.6 Hz), 160.2, (t,
compound 2a (46 mg, 0.050 mmol, 64%). ½a D20
ꢃ 19:7 (c 1.2, CHCl3);
ꢂ
1H NMR (400 MHz, CDCl3) d 8.15 (t, JHH = 1.4 Hz, 1H), 7.95–7.91 (m,
2H), 7.41–7.08 (m, 19H), 6.50–6.37 (m, 3H), 5.28–5.18 (m, 1H), 4.59
(d, JHH = 2.4 Hz, 2H), 4.41–4.27 (m, 2H), 4.23 (dd, JHH = 5.6, 8.8 Hz,
2H), 4.16 (dd, JHH = 5.4, 14.6 Hz, 1H), 4.08–3.95 (m, 3H), 3.27 (s, 3H),
2.80 (s, 3H), 2.33–2.21 (m, 1H), 2.20–2.08 (m, 2H), 1.98–1.89 (m,
1H), 1.55 (d, JHH = 6.9 Hz, 3H), 0.91 (d, JHH = 6.8 Hz, 3H), 0.84 (d,
JHH = 6.8 Hz, 3H); 13C NMR (100.5 MHz, CDCl3) d 173.2, 171.0, 166.5,
2
3JCF = 13.6 Hz), 138.4, 128.8, 127.9, 127.6, 98.6 (d, JCF = 29.1 Hz,
2
2C), 97.1 (t, JCF = 25.7 Hz), 78.6, 71.5, 70.7, 69.0, 68.0, 64.7, 59.1,
58.7, 43.6, 36.6, 30.0, 19.6, 17.6; MS (ESI) m/z 586.0 ([M+Na]+ calcd
þ
for C27H35F2N5NaO6 586.2).
1
1
165.0, 163.9 (d, JCF = 246.4 Hz), 163.7 (d, JCF = 246.4 Hz), 160.4 (t,
3JCF = 13.7 Hz), 143.1, 142.2, 138.2, 136.7, 136.0, 135.6, 129.0, 128.8,
128.7, 128.7, 128.1, 127.9, 127.6, 127.5, 126.4, 124.5, 98.7 (d,
2JCF = 28.8 Hz, 2C), 96.9 (t, 2JCF = 25.9 Hz), 77.0, 72.9, 67.3, 66.3, 58.3,
53.6, 50.0, 43.6, 36.0, 35.6, 30.2, 21.8, 19.7, 17.7; HRMS (ESI) m/z
950.3541 ([M+Na]+ calcd for C49H55F2N5NaO9S+ 950.3581). LC–MS
purity system A: tR = 5.19 min, 99%; system B: tR = 4.87 min, 100%.
4.6.20. 2-Allyloxy-5-azido-6-(3,5-difluoro-phenoxy)-4-hydroxy-
hexanoid acid (1-benzylcarbamoyl-2-methyl-propyl)-amide
(7d)
Compound 7d (39 mg, 0.071 mmol, 77%) was synthesized from
6d (31 mg, 0.092 mmol) according to the method for the prepara-
tion of 7a.
1H NMR (400 MHz, CDCl3) d 7.35–7.22 (m, 5H), 7.10 (d,
JHH = 8.8 Hz, 1H), 7.02 (t, JHH = 5.4 Hz, 1H), 6.50–6.39 (m, 3H),
5.97–5.86 (m, 1H), 5.38–5.26 (m, 2H), 4.42 (d, JHH = 5.5 Hz, 2H),
4.32 (dd, JHH = 5.3 Hz, 1H), 4.13–4.02 (m, 4H), 3.97 (t, JHH = 8.8 Hz,
2H), 3.70–3.63 (m, 1H), 2.82 (d, JHH = 7.3 Hz, 1H), 2.45–2.37 (m,
1H), 2.22–2.14 (m, 1H), 1.96–1.88 (m, 1H), 0.98 (d, JHH = 6.8 Hz,
3H), 0.94 (d, JHH = 6.8 Hz, 3H); 13C NMR (125.5 MHz, CDCl3) d
4.6.23. N-[(1S,2S,4R)-4-((S)-1-Benzylcarbamoyl-2-methyl-
propylcarbamoyl)-1-(3,5-difluoro-phenoxymethyl)-4-ethyl-2-
hydroxy-butyl]-5-(methanesulfonyl-methyl-amino)-N0-((R)-1-
phenyl-ethyl)-isophthalamide (2b)
Compound 2b (36 mg, 0.04 mmol, 67%) was synthesized from 7b
(67 mg, 0.13 mmol) that was reduced and then coupled to 5-(meth-
anesulfonyl-methyl-amino)-N0-(1-phenyl-ethyl)-isophthalacid
(24 mg, 0.06 mmol), according to the method for the preparation of
1
1
172.6, 170.9, 163.9 (d, JCF = 246.9 Hz), 163.8 (d, JCF = 246.9 Hz),
3
160.1 (t, JCF = 13.7 Hz), 138.3, 133.2, 128.8, 127.9, 127.6, 118.8,
2
2
98.7 (d, JCF = 28.9 Hz, 2C), 97.2 (t, JCF = 25.8 Hz), 77.1, 71.8, 69.0,
67.5, 64.8, 58.3, 43.7, 36.2, 30.0, 19.8, 17.6; MS (ESI) m/z 568.3
2a. ½a 2D0
ꢂ
ꢃ 19:8 (c 1.1, CHCl3); 1H NMR (400 MHz, CDCl3) d 8.15 (t,
JHH = 1.5 Hz, 1H), 7.96–7.93 (m, 2H), 7.35–7.09 (m, 14H), 7.04 (d,
JHH = 7.7 Hz, 1H), 6.51–6.37 (m, 3H), 5.29–5.20 (m, 1H), 4.42–4.31
(m, 2H), 4.28–4.17 (m, 3H), 4.07–4.02 (m, 2H), 3.92 (t, JHH = 5.1 Hz,
1H), 3.65–3.54 (m, 2H), 3.30 (s, 3H), 2.82 (s, 3H), 2.33–2.23 (m,
1H), 2.17–2.08 (m, 1H), 1.97–1.89 (m, 1H), 1.58 (d, JHH = 7.0 Hz,
3H), 1.27 (t, JHH = 7.0 Hz, 3H), 0.94 (d, JHH = 6.8 Hz, 3H), 0.90 (d,
JHH = 6.8 Hz, 3H); 13C NMR (100.5 MHz, CDCl3) d 173.4, 171.0,
([M+Na]+ calcd for C27H33F2N5NaO5 568.2).
þ
4.6.21. 5-Azido-2-cyclopropylmethoxy-6-(3,5-difluoro-
phenoxy)-4-hydroxy-hexanoid acid (1-benzylcarbamoyl-2-
methyl-propyl)-amide (7e)
Compound 7e (60 mg, 0.107 mmol, 97%) was synthesized from
6e (39 mg, 0.110 mmol) according to the method for the prepara-
tion of 7a. 1H NMR (400 MHz, CDCl3) d 7.36–7.20 (m, 5H), 7.15
(d, JHH = 8.8 Hz, 1H), 6.96 (t, JHH = 5.5 Hz, 1H), 6.51–6.37 (m, 3H),
4.42 (d, JHH = 5.7 Hz, 2H), 4.33–4.26 (m, 1H), 4.09–3.93 (m, 4H),
3.69–3.62 (m, 1H), 3.40 (d, JHH = 6.9 Hz, 2H), 2.85 (d, JHH = 6.9 Hz,
1H), 2.45–2.34 (m, 1H), 2.20–2.10 (m 1H), 1.95–1.87 (m, 1H),
1.13–1.04 (m, 1H), 0.98 (d, JHH = 6.9 Hz, 3H), 0.94 (d, JHH = 6.9 Hz,
1
1
166.4, 164.9, 163.9 (d, JCF = 246.4 Hz), 163.7 (d, JCF = 246.4 Hz),
3
160.4 (t, JCF = 13.7 Hz), 143.0, 142.3, 138.2, 136.0, 135.6, 128.9,
128.7, 128.0, 127.9, 127.7, 127.6, 126.4, 124.5, 98.7 (d, 2JCF = 28.6 Hz,
2
2C), 96.9 (t, JCF = 25.9 Hz), 77.9, 67.2, 66.7, 66.5, 58.3, 53.6, 50.0,
43.6, 38.1, 36.1, 35.6, 30.3, 21.9, 19.7, 17.8, 15.6; HRMS (ESI) m/z
888.3396 ([M+Na]+ calcd for C44H53F2N5NaO9S+ 888.3424). LC–MS
purity system A: tR = 4.89 min, 97%; system B: tR = 4.59 min, 99%.
3H), 0.64–0.57 (m, 2H), 0.27–0.21 (m, 2H); 13C NMR (100.5 MHz,
1
CDCl3)
d
172.8, 170.9, 163.9 (d, JCF = 246.8 Hz), 163.7 (d,
4.6.24. N-[(1S,2S,4R)-4-((S)-1-Benzylcarbamoyl-2-methyl-
propylcarbamoyl)-1-(3,5-difluoro-phenoxymethyl)-2-hydroxy-
4-(2-methoxy-ethoxy)-butyl]-5-(methanesulfonyl-methyl-
amino)-N0-((R)-1-phenyl-ethyl)-isophthalamide (2c)
Compound 2c (42 mg, 0.05 mmol, 97%) was synthesized from
7c (54 mg, 0.10 mmol) that was reduced and then coupled to
1JCF = 246.8 Hz), 160.1 (t, JCF = 13.7 Hz), 138.3, 128.8, 127.9,
127.6, 98.6 (d, JCF = 28.4 Hz, 2C), 97.2 (d, JCF = 25.7 Hz), 77.8,
3
2
2
75.4, 68.9, 67.9, 64.7, 58.3, 43.7, 36.2, 30.0, 19.7, 17.7, 10.8, 3.4,
3.3; MS (ESI) m/z 582.2 ([M+Na]+ calcd for C28H35F2N5NaO5
þ
582.3).

1720
C. Björklund et al. / Bioorg. Med. Chem. 18 (2010) 1711–1723
5-(methanesulfonyl-methyl-amino)-N0-(1-phenyl-ethyl)-isoph-
NMR (400 MHz, CDCl3) d 8.14–8.12 (m, 1H), 7.96–7.90 (m, 2H),
7.40–7.06 (m, 13H), 6.48–6.38 (m, 3H), 5.28–5.20 (m, 1H), 4.40–
4.30 (m, 2H), 4.28–4.16 (m, 3H), 4.08–3.98 (m, 2H), 3.90 (t,
JHH = 5.0 Hz, 1H), 3.82–3.74 (m, 1H), 3.56–3.43 (m, 2H), 3.28 (s,
3H), 2.81 (s, 3H), 2.32–2.22 (m, 1H), 2.16–2.04 (m, 2H), 1.98–1.90
(m, 1H), 1.69–1.59 (m, 2H), 1.57 (d, JHH = 6.9 Hz, 3H), 1.00–0.87
(m, 9H); 13C NMR (100.5 MHz, CDCl3) d 173.3, 171.0, 166.5, 165.1,
thalacid (18 mg, 0.05 mmol), according to the method for the prep-
aration of 2a. ½a D20
ꢂ
ꢃ 14:7 (c 1.3, CHCl3); 1H NMR (400 MHz, CDCl3)
d 8.16 (t, JHH = 1.4 Hz, 1H), 7.96–7.93 (m, 2H), 7.53 (d, JHH = 8.6 Hz,
1H), 7.37–7.09 (m, 14H), 6.50–6.37 (m, 3H), 5.31–5.21 (m, 1H),
4.43–4.32 (m, 2H), 4.29–4.17 (m, 3H), 4.07–4.02 (m, 2H), 3.95 (t,
JHH = 5.2 Hz, 1H), 3.76–3.70 (m, 1H), 3.68–3.49 (m, 3H), 3.37 (s,
3H), 3.30 (s, 3H), 2.82 (s, 3H), 2.30–2.19 (m, 1H), 2.16–2.05 (m,
1H), 1.98–1.89 (m, 1H), 1.58 (d, JHH = 7.0 Hz, 3H), 0.94 (d,
JHH = 6.9 Hz, 3H), 0.92 (d, JHH = 6.9 Hz, 3H); 13C NMR (100.5 MHz,
1
1
163.9 (d, JCF = 246.5 Hz), 163.7 (d, JCF = 246.5 Hz), 160.4 (t,
3JCF = 13.8 Hz), 143.1, 142.3, 138.2, 136.0, 135.6, 128.9, 128.7,
128.0, 127.9, 127.9, 127.6, 127.6, 126.5, 126.4, 124.4, 98.7 (d,
1
2
CDCl3) d 173.4, 171.1, 166.5, 165.0, 163.9 (d, JCF = 246.3 Hz),
2JCF = 28.2 Hz, 2C), 96.9 (t, JCF = 25.9 Hz), 78.0, 72.8, 67.2, 66.5,
1
3
163.7 (d, JCF = 246.3 Hz), 160.5 (t, JCF = 13.7 Hz), 143.1, 142.3,
58.2, 53.6, 50.0, 43.6, 38.0, 36.0, 35.7, 30.4, 29.8, 23.2, 21.8, 19.7,
17.7, 10.8; HRMS (ESI) m/z 880.3749 ([M+H]+ calcd for
C45H56F2N5O9S+ 880.3761). LC–MS purity system A: tR = 5.10 min,
99%; system B: tR = 4.79 min, 99%.
138.2, 136.0, 135.7, 128.9, 128.7, 128.0, 128.0, 127.9, 127.6,
2
127.5, 126.4, 124.5, 98.7 (d, JCF = 28.6 Hz, 2C), 96.8 (t,
2JCF = 25.9 Hz), 78.4, 77.4, 71.5, 70.6, 67.3, 66.7, 59.1, 58.8, 53.6,
50.0, 43.6, 38.1, 36.6, 35.7, 30.2, 21.8, 19.5, 17.7; HRMS (ESI) m/z
918.3523 ([M+Na]+ calcd for C45H55F2N5NaO9S+ 918.3530). LC–
MS purity system A: tR = 4.80 min, 95%; system B: tR = 4.51 min,
96%.
4.6.27. N-[(1S,2S,4R)-4-((S)-1-Benzylcarbamoyl-2-methyl-
propylcarbamoyl)-4-cyclopropylmethoxy-1-(3,5-difluoro-
phenoxymethyl)-2-hydroxy-butyl]-5-(methanesulfonyl-
methyl-amino)-N0-((R)-1-phenyl-ethyl)-isophthalamide (2f)
Compound 2f (27 mg, 0.03 mmol, 43%) was synthesized from 7e
(39 mg, 0.07 mmol) that was reduced and then coupled to 5-
(methanesulfonyl-methyl-amino)-N0-(1-phenyl-ethyl)-isophthal-
acid (34 mg, 0.09 mmol), according to the method for the prepara-
4.6.25. N-[(1S,2S,4R)-4-((S)-1-Benzylcarbamoyl-2-methyl-
propylcarbamoyl)-4-allyloxy-1-(3,5-difluoro-phenoxymethyl)-
2-hydroxy-butyl]-5-(methanesulfonyl-methyl-amino)-N0-((R)-
1-phenyl-ethyl)-isophthalamide (2d)
Compound 2d (18 mg, 0.02 mmol, 58%) was synthesized from
7d (19 mg, 0.04 mmol) that was reduced and then coupled to 5-
(methanesulfonyl-methyl-amino)-N0-(1-phenyl-ethyl)-isophthal-
acid (13 mg, 0.04 mmol), according to the method for the prepara-
tion of 2a. ½a 2D0
ꢂ
ꢃ 13:6 (c 1.0, MeOH); 1H NMR (400 MHz, MeOD-d4)
d 8.93 (d, JHH = 7.8 Hz, 1H), 8.60 (t, JHH = 6.0 Hz, 1H), 8.36 (d,
JHH = 8.7 Hz, 1H), 8.30–8.28 (t, JHH = 1.5 Hz, 1H), 8.10–8.07 (m,
1H), 8.05–8.03 (m, 1H), 7.71 (d, JHH = 8.7 Hz, 1H), 7.43–7.13 (m,
10H), 6.66–6.47 (m, 3H), 5.30–5.20 (m, 3H), 4.64–4.54 (m, 1H),
4.37–4.32 (m, 2H), 4.31–4.11 (m, 5H), 3.43–3.30 (m, 1H), 3.36 (s,
3H), 2.95 (s, 3H), 2.13–2.01 (m, 1H), 2.01–1.91 (m, 3H), 1.58 (d,
JHH = 7.0 Hz, 3H), 1.16–0.98 (m, 1H), 0.91 (d, JHH = 6.9 Hz, 3H),
0.87 (d, JHH = 6.9 Hz, 3H) 0.58–0.47 (m, 2H), 0.28–0.18 (m, 2H);
13C NMR (100.5 MHz, MeOD-d4) d 175.2, 173.2, 169.1, 167.7,
tion of 2a. ½a 2D0
ꢂ
þ 5:4 (c 1.3, CHCl3); 1H NMR (400 MHz, CDCl3) d
8.16–8.15 (m, 1H), 7.94–7.92 (m, 2H), 7.66–7.59 (m, 1H), 7.58–
7.51 (m, 1H), 7.48–7.41 (m, 1H), 7.34–7.05 (m, 12H), 6.48–6.37
(m, 3H), 5.95–5.84 (m, 1H), 5.34–5.19 (m, 3H), 4.39–4.30 (m,
2H), 4.30–4.16 (m, 3H), 4.10–3.97 (m, 5H), 3.27 (s, 3H), 2.81 (s,
3H), 2.33–2.21 (m, 1H), 2.20–2.06 (m, 1H), 1.97–1.90 (m, 1H),
1.56 (d, JHH = 7.0 Hz, 3H), 0.94 (d, JHH = 6.8 Hz, 3H), 0.89 (d,
JHH = 6.8 Hz, 3H); 13C NMR (100.5 MHz, CDCl3) d 173.3, 171.0,
1
1
165.3 (d, JCF = 245.1 Hz), 165.1 (d, JCF = 245.1 Hz), 162.2 (t,
3JCF = 13.8 Hz), 145.1, 143.8, 139.7, 137.4, 136.9, 129.6, 129.5,
129.4, 128.6, 128.3, 128.2, 127.3, 126.0, 99.6 (d, JCF = 28.7 Hz,
2C), 97.2 (t, JCF = 26.4 Hz), 78.3, 76.4, 68.6, 67.7, 59.8, 53.7, 51.0,
1
1
2
166.5, 165.0, 163.9 (d, JCF = 246.5 Hz), 163.7 (d, JCF = 246.5 Hz),
3
2
160.4 (t, JCF = 13.7 Hz), 143.1, 142.3, 138.2, 136.0, 135.6, 133.3,
132.3, 132.2, 128.8, 128.8, 128.7, 128.6, 128.1, 128.1, 127.9,
44.1, 38.4, 38.2, 35.9, 32.1, 22.2, 19.8, 18.6, 11.5, 3.8, 3.4; HRMS
(ESI) m/z 892.3778 ([M+H]+ calcd for C46H56F2N5O9S+ 892.3761).
LC–MS purity system A: tR = 5.23 min, 100%; system B:
tR = 4.82 min, 100%.
2
127.6, 127.6, 126.4, 124.5, 118.7, 98.7 (d, JCF = 28.2 Hz, 2C), 96.9
2
(t, JCF = 25.9 Hz) 77.0, 71.7, 67.2, 66.2, 58.3, 53.6, 50.0, 43.6, 38.1,
36.0, 35.7, 30.2, 21.9, 19.7, 17.7; HRMS (ESI) m/z 878.3580
([M+H]+ calcd for C45H54F2N5O9S+ 878.3605). LC–MS purity system
A: tR = 5.00 min, 99%; system B: tR = 4.69 min, 99%.
4.6.28. (2R,4S,5S)-5-Azido-6-(3,5-difluorophenoxy)-4-hydroxy-
2-methoxy-hexanoic acid ((S)-1-benzylcarbamoyl-2-methyl-
propyl)-amide (8)
4.6.26. N-[(1S,2S,4R)-4-((S)-1-Benzylcarbamoyl-2-methyl-
propylcarbamoyl)-1-(3,5-difluoro-phenoxymethyl)-2-hydroxy-
4-propyloxy-butyl]-5-(methanesulfonyl-methyl-amino)-N0-
((R)-1-phenyl-ethyl)-isophthalamide (2e)
Compound 8 was synthesized according to a protocol previ-
ously reported by our group.11
Azide 7d (18 mg, 0.034 mmol) was dissolved in MeOH (4 mL) and
a catalytical amount of Pd on carbon was added, the mixture was
then treated with hydrogen gas (1 atm) for 2 h. Then the solids were
filtered off and the filtrate was concentrated under vacuum. Without
further purification the formed amine was used in the next step. 5-
(Methanesulfonyl-methyl-amino)-N0-(1-phenyl-ethyl)-isophthal-
acid (13 mg, 0.034 mmol, 1 equiv), Py-BOP (18 mg, 0.034 mmol,
4.6.29. N-[(1S,4R)-4-((S)-1-Benzylcarbamoyl-2-methyl-
propylcarbamoyl)-1-(3,5-difluoro-phenoxymethyl)-4-methoxy-
butyl]-5-(methanesulfonyl-methyl-amino)-N0-((R)-1-phenyl-
ethyl)-isophthalamide (2g)
A solution of 8 (54 mg, 0.104 mmol) in dry dichloroethane
(5 mL) was refluxed. 1,10-Thiocarbonyldiimidazole (32 mg,
0.177 mmol, 1.7 equiv) was added to the solution and the refluxing
was continued for 2 h. The solvent was concentrated under vac-
uum. Without further purification the formed thiocarbonyl ester
was used in the next step. To a refluxing solution of Bu3SnH
1 equiv) and DIPEA (6 lL, 0.034 mmol, 1 equiv) were dissolved in
DCM (2 mL). The mixture was stirred at room temperature for
30 min before the amine (ꢀ1 equiv) from the previous reaction dis-
solved in DCM (2 mL) and DIPEA (6
lL, 0.034 mmol, 1 equiv) were
(55 lL, 0.207 mmol, 2 equiv) and a catalytic amount of AIBN in tol-
added. After complete reaction the mixture were washed with NaH-
CO3 (1ꢁ 10 mL) and brine (1ꢁ 10 mL). The water phase was ex-
tracted with DCM (2ꢁ 10 mL). The organic layers were combined
and dried over Na2SO4, concentrated under vacuum and purified
by column chromatography (toluene/ethyl acetate 1:1) to give com-
uene (10 mL), a solution of the previously prepared thiocarbonyl
ester (0.104 mmol, 1 equiv) in toluene (3 mL) was added drop wise.
The reaction mixture was refluxed for 2 h. The solvent was concen-
trated under vacuum. Without further purification the formed
amine was used in the next step. 5-(Methanesulfonyl-methyl-
amino)-N0-(1-phenyl-ethyl)-isophthalacid (40 mg, 0.106 mmol,
pound 2e (26 mg, 0.030 mmol, 88%). ½a D20
ꢂ
þ 6:0 (c 0.8, CHCl3); 1H

C. Björklund et al. / Bioorg. Med. Chem. 18 (2010) 1711–1723
1721
1 equiv), Py-BOP (56 mg, 0.106 mmol, 1 equiv) and DIPEA (19
l
L,
Without further purification the formed amine was used in the
next step.
5-(Methanesulfonyl-methyl-amino)-N0-(1-phenyl-
ethyl)-isophthalacid (16 mg, 0.042 mmol, 1 equiv), Py-BOP
(22 mg, 0.042 mmol, 1 equiv) and DIPEA (7.5 L, 0.042 mmol,
0.109 mmol, 1 equiv) were dissolved in DCM (2 mL). The mixture
was stirred at room temperature for 30 min before the amine
(ꢀ1 equiv) from the previous reaction dissolved in DCM (2 mL)
l
and DIPEA (23
l
L, 0.138 mmol, 1.3 equiv) were added. After com-
1 equiv) were dissolved in DCM (2 mL). The mixture was stirred
plete reaction the mixture were washed with NaHCO3 (1ꢁ
10 mL) and brine (1ꢁ 10 mL). The water phase was extracted with
DCM (2ꢁ 10 mL). The organic layers were combined and dried over
Na2SO4, concentrated under vacuum and purified by column chro-
matography (toluene/ethyl acetate 1:1) to give compound 2g
at room temperature for 30 min before the amine (ꢀ1 equiv) from
the previous reaction dissolved in DCM (2 mL) and DIPEA (7.5 lL,
0.042 mmol, 1 equiv) were added. After complete reaction the mix-
ture were washed with NaHCO3 (1ꢁ 10 mL) and brine (1ꢁ 10 mL).
The water phase was extracted with DCM (2ꢁ 10 mL). The organic
layers were combined and dried over Na2SO4, concentrated under
vacuum and purified by column chromatography (toluene/ethyl
acetate 1:1) to give compound 2h (27 mg, 0.032 mmol, 75%).
(6 mg, 0.007 mmol, 7%).
(400 MHz, MeOD-d4)
½
a 2D0
ꢂ
ꢃ 4:2 (c 0.4, MeOH); 1H NMR
d
8.92 (d, JHH = 7.8 Hz, 1H), 8.60 (t,
JHH = 5.9 Hz, 1H), 8.53 (d, JHH = 8.4 Hz, 1H), 8.22 (t, JHH = 1.5 Hz,
1H), 8.02 (d, JHH = 1.5 Hz, 2H), 7.80 (d, JHH = 8.7 Hz, 1H), 7.43–
7.39 (m, 2H), 7.35–7.29 (m, 2H), 7.28–7.13 (m, 6H), 6.64–6.47
(m, 3H), 5.29–5.20 (m, 1H), 4.50–4.40 (m, 1H), 4.38–4.27 (m,
2H), 4.23–4.17 (m, 1H), 4.12–4.03 (m, 2H), 3.41 (s, 3H), 3.35 (s,
3H), 2.94 (s, 3H), 2.11–2.01 (m, 1H), 1.95–1.68 (m, 4H), 1.58 (d,
JHH = 7.1 Hz, 3H), 0.93 (d, JHH = 6.8 Hz, 3H), 0.93 (d, JHH = 6.8 Hz,
3H); 13C NMR (100.5 MHz, MeOD-d4) d 174.9, 173.3, 168.8, 167.8,
½
a 2D0
ꢂ
þ 5:1 (c 1.0, MeOH); 1H NMR (400 MHz, MeOD-d4) d 8.20 (t,
JHH = 1.6 Hz, 1H), 8.02–7.98 (m, 2H), 7.42–7.38 (m, 2H), 7.35–7.29
(m, 2H), 7.28–7.08 (m, 7H), 6.63–6.46 (m, 3H), 5.27–5.20 (m,
1H), 4.44–4.37 (m, 1H), 4.35–4.25 (m, 4H), 4.20 (d, JHH = 7.6 Hz,
1H), 4.13–4.06 (m, 1H), 3.99 (dd, JHH = 2.8, 10.2 Hz, 1H), 3.43 (s,
3H), 3.33 (s, 3H), 2.92 (s, 3H), 2.32 (s, 1H), 2.13–2.02 (m, 1H),
1.99–1.90 (m, 1H), 1.88–1.78 (m, 1H), 1.57 (d, JHH = 7.1 Hz, 3H),
0.96–0.93 (m, 6H); 13C NMR (100.5 MHz, MeOD-d4) d 175.4,
1
1
165.3 (d, JCF = 244.8 Hz), 165.1 (d, JCF = 244.8 Hz), 162.4 (t,
3JCF = 13.6 Hz), 145.1, 143.8, 139.6, 137.4, 137.2, 129.6, 129.5,
129.3, 129.2, 128.6, 128.3, 128.2, 127.3, 125.9, 99.6 (d,
1
173.3, 168.8, 167.8, 165.2 (d, JCF = 244.8 Hz), 165.1 (d,
1JCF = 244.8 Hz), 162.5 (t, JCF = 13.9 Hz), 145.1, 143.7, 139.6,
3
2
2JCF = 28.8 Hz, 2C), 97.2 (t, JCF = 26.6 Hz), 82.5, 71.3, 60.0, 58.6,
137.4, 137.2, 129.9, 129.6, 129.5, 129.4, 129.2, 129.2, 128.6,
2
51.0, 50.5, 44.1, 38.3, 35.9, 32.1, 30.1, 27.3, 22.2, 19.8, 18.9; HRMS
(ESI) m/z 836.3485 ([M+Na]+ calcd for C43H52F2N5O8S+ 836.3499).
LC–MS purity system A: tR = 5.08 min, 99%; system B: tR = 4.66 min,
100%.
128.3, 128.2, 127.3, 126.3, 125.9, 99.6 (d, JCF = 28.7 Hz, 2C), 97.1
2
(t, JCF = 26.3 Hz), 80.3, 68.7, 68.0, 60.0, 59.1, 55.7, 51.0, 44.0,
39.1, 38.3, 35.9, 32.1, 22.2, 19.8, 18.8; HRMS (ESI) m/z 852.3433
([M+H]+ calcd for C43H52F2N5O9S+ 852.3448). LC–MS purity system
A: tR = 4.45 min, 99%; system B: tR = 4.12 min, 100%.
4.6.30. (2R,4R,5S)-5-Azido-6-(3,5-difluoro-phenoxy)-4-
hydroxy-2-methoxy-hexanoic acid ((S)-1-benzylcarbamoyl-2-
methyl-propyl)-amide (9)
To a stirred solution of 8 (226 mg, 0.435 mmol) in THF, p-nitro-
benzoic acid (131 mg, 0.783 mmol, 1.8 equiv) and triphenyl phos-
phine (194 mg, 0.74 mmol, 1.7 equiv) were added. The mixture
was cooled to 0 °C (ice/water). Diisopropyl azodicarboxylate
4.6.32. [(1S,2R,4R)-4-((S)-1-Benzylcarbamoyl-2-methyl-
propylcarbamoyl)-1-(3,5-difluoro-phenoxymethyl)-2-hydroxy-
4-methoxy-butyl]-carbamic acid tert-butyl ester (10)
Azide 9 (53 mg, 0.102 mmol) was dissolved in MeOH (8 mL) and
a catalytical amount of Pd on carbon was added, the mixture was
then treated with hydrogen gas (1 atm) for 2 h. Then the solids
were filtered off and the filtrate was concentrated under vacuum.
To a mixture of the formed amine dissolved in THF/H2O 1:1
(4 mL), 1 M NaOH in water (0.2 mL, 0.204 mmol, 2 equiv) and di-
tert-butyl dicarbonate (44.5 mg, 0.204 mmol, 2 equiv) were added.
The reaction was stirred in room temperature for 4 h. The volatile
solvents were evaporated off and the residue was neutralized using
1 M HCl and then extracted with DCM (3ꢁ 7 mL). The organic layer
was dried over Na2SO4, concentrated under vacuum and purified
by column chromatography (toluene/ethyl acetate 2:1) to give
compound 10 (58 mg, 0.098 mmol, 96%). 1H NMR (400 MHz,
CDCl3) d 7.33–7.17 (m, 6H), 6.80–6.70 (m, 1H), 6.46–6.34 (m,
3H), 5.08 (d, JHH = 8.0 Hz, 1H), 4.48–4.30 (m, 2H), 4.28–4.20 (m,
1H), 4.17–4.00 (m, 2H), 4.00–3.78 (m, 4H), 3.43 (s, 3H), 2.26–2.15
(m, 1H), 2.05–1.81 (m, 2H), 1.42 (s, 9H), 0.96 (d, JHH = 6.8 Hz, 3H),
0.93 (d, JHH = 6.8 Hz, 3H); 13C NMR (100.5 MHz, CDCl3) d 173.3,
(146 lL, 0.74, 1.7 equiv) was added drop wise to the solution.
The reaction was stirred in room temperature over night. The reac-
tion mixture was concentrated and the residue was filtered
through a short silica column, the filtrate was concentrated and
the formed ester was dissolved in MeOH (10 mL), to this mixture
1 M NaOMe in MeOH (1 mL) was added. The reaction was stirred
in room temperature for 1 h and then Dowex H+ was added to neu-
tralize the mixture. The solids were filtered off and the filtrate was
concentrated under vacuum. The residue was purified by column
chromatography (toluene/ethyl acetate 5:1) to give compound 9
(53 mg, 0.102 mmol, 23%). 1H NMR (400 MHz, CDCl3) d 7.40–7.17
(m, 6H), 6.61 (t, JHH = 5.7 Hz, 1H), 6.49–6.39 (m, 3H), 4.51–4.36
(m, 2H), 4.28–4.18 (m, 2H), 4.01–3.94 (m, 1H), 3.94–3.88 (m,
1H), 3.83–3.75 (m, 1H), 3.68–3.61 (m, 1H), 3.47 (s, 3H), 2.29–
2.17 (m, 1H), 2.09–2.00 (m, 1H), 1.93–1.82 (m, 1H), 0.98 (d,
JHH = 6.8 Hz, 3H), 0.96 (d, JHH = 6.8 Hz, 3H); 13C NMR (100.5 MHz,
170.8, 163.8 (d, JCF = 246.1 Hz), 163.7 (d, JCF = 246.1 Hz), 160.6
1
1
1
3
CDCl3)
d
173.3, 170.7, 163.9 (d, JCF = 246.7 Hz), 163.7 (d,
(t, JCF = 13.7 Hz), 155.9, 146.9, 138.1, 128.8, 127.8, 127.6, 98.6 (d,
1JCF = 246.7 Hz), 160.2 (t, JCF = 13.7 Hz), 137.9, 128.9, 127.9,
127.8, 98.7 (d, JCF = 28.5 Hz, 2C), 97.1 (t, JCF = 26.0 Hz), 80.3,
2JCF = 28.4 Hz, 2C), 96.8 (t, JCF = 25.8 Hz), 85.3, 80.5, 80.1, 68.9,
3
2
2
2
67.6, 58.8, 58.5, 54.3, 43.6, 36.6, 30.8, 28.5, 27.5, 19.6, 18.3; MS
(ESI) m/z 616.3 ([M+Na]+ calcd for C30H41F2N3NaO7 616.3).
þ
69.0, 68.5, 65.0, 58.9, 58.7, 43.7, 36.0, 30.8, 19.6, 18.3; MS (ESI)
m/z 520.2 ([M+H]+ calcd for C25H32F2N5O5 520.2).
þ
4.6.33. [(1S,2S,4R)-2-Azido-4-((S)-1-benzylcarbamoyl-2-methyl-
propylcarbamoyl)-1-(3,5-difluoro-phenoxymethyl)-4-methoxy-
butyl]-carbamic acid tert-butyl ester (11)
To a stirred solution of 10 (58 mg, 0.098 mmol) in pyridine
(5 mL), methane sulfonyl chloride (15 lL, 0.195, 2 equiv) was
4.6.31. N-[(1S,2R,4R)-4-((S)-1-Benzylcarbamoyl-2-methyl-
propylcarbamoyl)-1-(3,5-difluoro-phenoxymethyl)-2-hydroxy-
4-methoxy-butyl]-5-(methanesulfonyl-methyl-amino)-N0-((R)-
1-phenyl-ethyl)-isophthalamide (2h)
Azide 9 (22 mg, 0.042 mmol) was dissolved in MeOH (5 mL) and
a catalytical amount of Pd on carbon was added, the mixture was
then treated with hydrogen gas (1 atm) for 2 h. Then the solids
were filtered off and the filtrate was concentrated under vacuum.
added drop wise at 0 °C. The reaction was stirred in room temper-
ature for 1 h. The reaction mixture was concentrated under vac-
uum, dissolved in toluene and concentrated again. The formed
mesylate was dissolved in DMF (4 mL) and treated with sodium

1722
C. Björklund et al. / Bioorg. Med. Chem. 18 (2010) 1711–1723
azide (95.3 mg, 1.47 mmol, 15 equiv). The mixture was stirred at
60 °C over night. The reaction mixture was concentrated under
vacuum. The residue was dissolved in diethyl ether (20 mL) and
washed with H2O (1ꢁ 10 mL), Brine (1ꢁ 10 mL) and H2O (1ꢁ
10 mL) again. The organic layer was dried over Na2SO4, concen-
trated under vacuum and purified by column chromatography (tol-
uene/ethyl acetate 9:1) to give compound 11 (15 mg, 0.024 mmol,
25%). 1H NMR (400 MHz, MeOD-d4) d 7.33–7.26 (m, 4H), 7.25–7.18
(m, 2H), 7.17–7.09 (m, 1H), 6.89 (d, JHH = 9.0 Hz, 1H), 6.65–6.47 (m,
3H), 4.43–4.32 (m, 2H), 4.22 (d, JHH = 7.7 Hz, 1H), 4.12–4.00 (m,
3H), 3.96–3.88 (m, 2H), 3.41 (s, 3H), 2.14–1.93 (m, 3H), 1.46 (s,
9H), 0.95 (d, JHH = 6.8 Hz, 3H), 0.95 (d, JHH = 6.8 Hz, 3H); 13C NMR
The residue was purified by column chromatography (toluene/
ethyl acetate 1:6?ethyl acetate/ammonia 1:0.01) to give com-
pound 2j (14 mg, 0.016 mmol, 85%). ½a D20
ꢂ
ꢃ 1:9 (c 0.9, MeOH); 1H
NMR (500 MHz, MeOD-d4) d 8.28 (t, JHH = 1.6 Hz, 1H), 8.09–8.08
(m, 1H), 8.05–8.03 (m, 1H), 7.43–7.39 (m, 2H), 7.35–7.30 (m,
2H), 7.29–7.13 (m, 6H), 6.65–6.49 (m, 3H), 5.28–5.22 (m, 1H),
4.61–4.55 (m, 1H), 4.39–4-30 (m, 2H), 4.27–4.20 (m, 2H), 4.18 (d,
JHH = 7.4 Hz, 1H), 4.03–3.97 (m, 1H), 3.40 (s, 3H), 3.36 (s, 3H),
2.96 (s, 3H), 2.12–2.03 (m, 1H), 1.99–1.95 (m, 1H), 1.86–1.77 (m,
1H), 1.58 (d, JHH = 7.1 Hz, 3H), 0.91 (d, JHH = 6.8 Hz, 3H), 0.90 (d,
JHH = 6.8 Hz, 3H); 13C NMR (125.8 MHz, MeOD-d4) d 174.7, 173.3
1
1
169.1, 167.8, 165.2 (d, JCF = 244.6 Hz), 165.1 (d, JCF = 244.6 Hz),
1
3
(100.5 MHz, MeOD-d4) d 174.1, 173.2, 165.2 (d, JCF = 244.9 Hz),
162.1 (t, JCF = 13.9 Hz), 145.1, 143.8, 139.7, 137.5, 136.9, 129.6,
1
3
165.1 (d, JCF = 244.9 Hz), 162.0 (t, JCF = 13.8 Hz), 158.2, 139.7,
129.5, 129.5, 129.5, 129.4, 128.6, 128.3, 128.2, 127.3, 126.1, 99.7
2
2
2
129.5, 128.7, 128.3, 99.7 (d, JCF = 28.7 Hz, 2C), 97.4 (t,
(d, JCF = 29.1 Hz, 2C), 97.3 (t, JCF = 26.5 Hz), 81.1, 69.2, 60.0, 58.6,
53.7, 51.0, 50.3, 44.0, 38.4, 38.1, 36.0, 32.0, 22.1, 22.0, 19.8, 18.8;
HRMS (ESI) m/z 851.3612 ([M+H]+ calcd for C43H52F2N6O8S+
851.3608). LC–MS Purity system A: tR = 4.38 min, 97%; System B:
tR = 4.09 min, 97%.
2JCF = 26.3 Hz), 80.7, 80.3, 69.1, 60.5, 60.0, 58.7, 53.8, 44.1, 34.9,
32.2, 28.7, 19.8, 18.9; MS (ESI) m/z 641.3 ([M+Na]+ calcd for
þ
C30H40F2N6O6 641.3).
4.6.34. N-[(1S,2S,4R)-2-Azido-4-((S)-1-benzylcarbamoyl-2-
methyl-propylcarbamoyl)-1-(3,5-difluoro-phenoxymethyl)-4-
methoxy-butyl]-5-(methanesulfonyl-methyl-amino)-N0-((R)-1-
phenyl-ethyl)-isophthalamide (2i)
To a stirred solution of 11 (15 mg, 0.024 mmol), triethyl silane
(10 lL, 0.06 mmol, 2.5 equiv) in DCM (2 mL) trifluoroacetic acid
Acknowledgments
We gratefully thank the following personnel at Medivir AB:
Kurt Benkestock for conducting LC–MS purity measurements, Eliz-
abeth Hamelink and Ian Henderson for BACE-1 and Cath D enzyme
data. Finally, we would like to acknowledge Medivir AB for per-
forming the X-ray crystal studies and for financial support.
(1 mL) was added. After 15 min the mixture was concentrated un-
der vacuum, dissolved in toluene and concentrated again. The res-
idue was dissolved in DCM (10 mL), washed with NaHCO3 (2ꢁ
5 mL), dried over Na2SO4 and concentrated under vacuum. With-
out further purification the formed amine was used in the next
References and notes
5-(Methanesulfonyl-methyl-amino)-N0-(1-phenyl-ethyl)-
1. Glenner, G. G.; Wong, C. W. Biochem. Biophys. Res. Commun. 1984, 120, 885.
2. Goedert, M.; Wischik, C. M.; Crowther, R. A.; Walker, J. E.; Klug, A. Proc. Natl.
Acad. Sci. U.S.A. 1988, 85, 4051.
3. Selkoe, D. J. Nature 1999, 399, A23.
4. Hardy, J. A.; Higgins, G. A. Science 1992, 256, 184.
5. Hills, I. D.; Vacca, J. P. Curr. Opin. Drug Discovery Dev. 2007, 10, 383.
6. Shiuto, D.; Kasai, S.; Kimura, T.; Liu, P.; Hidaka, K.; Hamada, T.; Shibakawa, S.;
Hayashi, Y.; Hattori, C.; Szabó, B.; Ishiura, S.; Kiso, Y. Bioorg. Med. Chem. Lett.
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7. Ziora, Z.; Kimura, T.; Kiso, Y. Drugs Future 2006, 31, 53.
8. Stachel, S. J. Drug Dev. Res. 2009, 70, 101.
9. Ghosh, A. K.; Kumaragurubaran, N.; Hong, L.; Koelsch, G.; Tang, J. Curr.
Alzheimer Res. 2008, 5, 121.
10. Silvestri, R. Med. Res. Rev. 2009, 29, 295.
step.
isophthalacid (14 mg, 0.036 mmol, 1.5 equiv), Py-BOP (19 mg,
0.036 mmol, 1.5 equiv) and DIPEA (7 L, 0.036 mmol, 1.5 equiv)
l
were dissolved in DCM (1 mL). The mixture was stirred at room
temperature for 30 min before the amine (ꢀ1 equiv) from the pre-
vious reaction dissolved in DCM (2 mL) and DIPEA (5 lL,
0.024 mmol, 1 equiv) were added. After complete reaction the mix-
ture were washed with NaHCO3 (1ꢁ 10 mL) and brine (1ꢁ 10 mL).
The water phase was extracted with DCM (2ꢁ 10 mL). The organic
layers were combined and dried over Na2SO4, concentrated under
vacuum and purified by column chromatography (toluene/ethyl
acetate 1:1) to give compound 2i (18 mg, 0.021 mmol, 85%).
11. Björklund, C.; Oscarson, S.; Benkestock, K.; Borkakoti, N.; Jansson, K.; Lindberg,
J.; Hallberg, A.; Rosenquist, Å.; Samuelsson, B., J. Med. Chem., accepted for
publication.
½
a 2D0
ꢂ
ꢃ 4:9 (c 0.9, MeOH); 1H NMR (400 MHz, MeOD-d4) d 8.26 (t,
12. RCSB PDB (3IXJ).
JHH = 1.5 Hz, 1H), 8.06–8.05 (m, 1H), 8.05–8.03 (m, 1H), 7.42–7.38
(m, 2H), 7.35–7.29 (m, 2H), 7.28–7.16 (m, 6H), 6.67–6.50 (m,
3H), 5.28–5.21 (m, 1H), 4.77–4.71 (m, 1H), 4.40–4.30 (m, 2H),
4.26–4.19 (m, 3H), 4.11–4.05 (m, 1H), 4.05–4.00 (m, 1H), 3.43 (s,
3H), 3.36 (s, 3H), 2.95 (s, 3H), 2.13–2.00 (m, 3H), 1.58 (d,
JHH = 7.0 Hz, 3H), 0.92 (d, JHH = 6.8 Hz, 3H), 0.91 (d, JHH = 6.8 Hz,
3H); 13C NMR (100.5 MHz, MeOD-d4) d 174.1, 173.2, 169.2, 167.8,
13. Sauder, J. M.; Arthur, J. W.; Dunbrack, R. L., Jr. J. Mol. Biol. 2000, 300, 241.
14. Brady, S. F.; Singh, S.; Crouthamel, M.-C.; Holloway, M. K.; Coburn, C. A.;
Garsky, V. M.; Bogusky, M.; Pennington, M. W.; Vacca, J. P.; Hazuda, D.; Lai, M.-
T. Bioorg. Med. Chem. Lett. 2004, 14, 601.
15. La Regina, G.; Piscitelli, F.; Silvestri, R. J. Heterocycl. Chem. 2009, 46, 10.
16. Gou, T.; Hobbs, D. W. Curr. Med. Chem. 2006, 13, 1811.
17. Stachel, S. J.; Coburn, C. A.; Steele, T. G.; Jones, K. G.; Loutzenhiser, E. F.; Gregro,
A. R.; Rajapakse, H. A.; Lai, M.-T.; Crouthamel, M.-C.; Xu, M.; Tugusheva, K.;
Lineberger, J. E.; Pietrak, B. L.; Espeseth, A. S.; Shi, X.-P.; Chen-Dodson, E.;
Holloway, M. K.; Munshi, S.; Simon, A. J.; Kuo, L.; Vacca, J. P. J. Med. Chem. 2004,
47, 6447.
18. Schmidt, O. T.. In Methods in Carbohydrate Chemistry; Whistler, R. L., Woffrom, M.
L., BeMiller, J. N., Eds.; Academic Press: New York-London, 1963; Vol. II, p 318.
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S.; Hallberg, A.; Samuelsson, B. Bioorg. Med. Chem. 2003, 11, 1107.
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21. Johansson, P.-O.; Chen, Y.; Belfrage, A. K.; Blackman, M. J.; Kvarnström, I.;
Jansson, K.; Vrang, L.; Hamelink, E.; Hallberg, A.; Rosenquist, Å.; Samuelsson, B.
J. Med. Chem. 2004, 47, 3353.
1
1
165.3 (d, JCF = 244.9 Hz), 165.1 (d, JCF = 244.9 Hz), 161.9 (t,
3JCF = 13.8 Hz), 145.1, 143.8, 139.7, 137.5, 136.8, 129.6, 129.5,
2
129.3, 128.6, 128.3, 128.2, 127.3, 126.1, 99.7 (d, JCF = 28.7 Hz,
2C), 97.5 (t, JCF = 26.3 Hz), 80.1, 68.7, 60.5, 60.1, 58.7, 53.0, 51.0,
2
44.1, 38.3, 35.9, 35.2, 32.2, 22.2, 19.8, 18.9; HRMS (ESI) m/z
899.3324 ([M+Na]+ calcd for C43H50F2N8O8S+ 899.3333). LC–MS
purity system A: tR = 4.95 min, 99%; system B: tR = 4.68 min, 100%.
4.6.35. N-[(1S,2S,4R)-4-((S)-1-Benzylcarbamoyl-2-methyl-
propylcarbamoyl)-2-amino-1-(3,5-difluoro-phenoxymethyl)-4-
methoxy-butyl]-5-(methanesulfonyl-methyl-amino)-N0-((R)-1-
phenyl-ethyl)-isophthalamide (2j)
Azide 2i (17 mg, 0.019 mmol) was dissolved in MeOH (4 mL)
and a catalytical amount of Pd on carbon was added, the mixture
was then treated with hydrogen gas (1 atm) for 2 h. Then the solids
were filtered off and the filtrate was concentrated under vacuum.
22. Zuccarello, G.; Bouzide, A.; Kvarnström, I.; Niklasson, G.; Svensson, S. C. T.;
Brisander, M.; Danielsson, H.; Nillroth, U.; Karlén, A.; Hallberg, A.; Classon, B.;
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