Discovery of orally efficacious RORγt inverse agonists. Part 2: Design, synthesis, and biological evaluation of novel tetrahydroisoquinoline derivatives

Article abstract of DOI:10.1016/j.bmc.2017.12.008

A series of tetrahydroisoquinoline derivatives were designed, synthesized, and evaluated for their potential as novel orally efficacious retinoic acid receptor-related orphan receptor-gamma t (RORγt)inverse agonists for the treatment of Th17-driven autoimmune diseases. We carried out cyclization of the phenylglycinamide core by structure-based drug design and successfully identified a tetrahydroisoquinoline carboxylic acid derivative 14 with good biochemical binding and cellular reporter activity. Interestingly, the combination of a carboxylic acid tether and a central fused bicyclic ring was crucial for optimizing PK properties, and the compound 14 showed significantly improved PK profile. Successive optimization of the carboxylate tether led to the discovery of compound 15 with increased inverse agonistic activity and an excellent PK profile. Oral treatment of mice with compound 15 robustly and dose-dependently inhibited IL-17A production in an IL23-induced gene expression assay.

Full text of DOI:10.1016/j.bmc.2017.12.008

Accepted Manuscript
Discovery of orally efficacious RORγt inverse agonists. Part 2: Design, syn-
thesis, and biological evaluation of novel tetrahydroisoquinoline derivatives
Mitsunori Kono, Tsuneo Oda, Michiko Tawada, Takashi Imada, Yoshihiro
Banno, Naohiro Taya, Tetsuji Kawamoto, Hidekazu Tokuhara, Yoshihide
Tomata, Naoki Ishii, Atsuko Ochida, Yoshiyuki Fukase, Tomoya Yukawa, Shoji
Fukumoto, Hiroyuki Watanabe, Keiko Uga, Akira Shibata, Hideyuki
Nakagawa, Mikio Shirasaki, Yasushi Fujitani, Masashi Yamasaki, Junya Shirai,
Satoshi Yamamoto
PII:
S0968-0896(17)31988-0
https://doi.org/10.1016/j.bmc.2017.12.008
BMC 14113
DOI:
Reference:
Bioorganic & Medicinal Chemistry
To appear in:
Received Date:
Revised Date:
Accepted Date:
11 October 2017
30 November 2017
3 December 2017
Please cite this article as: Kono, M., Oda, T., Tawada, M., Imada, T., Banno, Y., Taya, N., Kawamoto, T., Tokuhara,
H., Tomata, Y., Ishii, N., Ochida, A., Fukase, Y., Yukawa, T., Fukumoto, S., Watanabe, H., Uga, K., Shibata, A.,
Nakagawa, H., Shirasaki, M., Fujitani, Y., Yamasaki, M., Shirai, J., Yamamoto, S., Discovery of orally efficacious
RORγt inverse agonists. Part 2: Design, synthesis, and biological evaluation of novel tetrahydroisoquinoline
derivatives, Bioorganic & Medicinal Chemistry (2017), doi: https://doi.org/10.1016/j.bmc.2017.12.008
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers
we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and
review of the resulting proof before it is published in its final form. Please note that during the production process
errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

Discovery of orally efficacious RORγt inverse agonists. Part 2: Design, synthesis, and
biological evaluation of novel tetrahydroisoquinoline derivatives
Mitsunori Kono,*a Tsuneo Oda,a Michiko Tawada,a Takashi Imada,a Yoshihiro Banno,a,b Naohiro
Taya,a,b Tetsuji Kawamoto,a,b Hidekazu Tokuhara,a,b Yoshihide Tomata,a Naoki Ishii,a Atsuko
Ochida,a Yoshiyuki Fukase,a,c Tomoya Yukawa,a Shoji Fukumoto,a,d Hiroyuki Watanabe,a Keiko
Uga,a Akira Shibata,a Hideyuki Nakagawa,a Mikio Shirasaki,a,b Yasushi Fujitani,a Masashi
Yamasaki,a,b Junya Shirai,a,e and Satoshi Yamamotoa
a Pharmaceutical Research Division, Takeda Pharmaceutical Company Limited, 26-1 Muraoka-
Higashi 2-chome, Fujisawa, Kanagawa 251-8555, Japan
ABSTRACT
A series of tetrahydroisoquinoline derivatives were designed, synthesized, and evaluated for their
potential as novel orally efficacious retinoic acid receptor-related orphan receptor-gamma t
(RORγt) inverse agonists for the treatment of Th17-driven autoimmune diseases. We carried out
cyclization of the phenylglycinamide core by structure-based drug design and successfully
identified a tetrahydroisoquinoline carboxylic acid derivative 14 with good biochemical binding
and cellular reporter activity. Interestingly, the combination of a carboxylic acid tether and a
central fused bicyclic ring was crucial for optimizing PK properties, and the compound 14

showed significantly improved PK profile. Successive optimization of the carboxylate tether led
to the discovery of compound 15 with increased inverse agonistic activity and an excellent PK
profile. Oral treatment of mice with compound 15 robustly and dose-dependently inhibited IL-
17A production in an IL23-induced gene expression assay.
KEYWORDS: Retinoic acid receptor-related orphan receptor-gamma t (RORγt), inverse agonist,
autoimmune disease, Th17, IL-17, tetrahydroisoquinoline
ABBREVIATIONS: ROR t, retinoic acid receptor-related orphan receptor-gamma t; EAE,
experimental autoimmune encephalomyelitis; BODIPY, boron-dipyrromethene; TR-FRET, time-
resolved fluorescence resonance energy transfer; SAR, structure-activity relationship; IFN- ,
interferon- ; PBS, phosphate buffered saline; SBDD, structure-based drug design; PK,
pharmacokinetics; PD, pharmacodynamics; AUC, area under the plasma concentration-time
curve; F, bioavailability
*Corresponding author. Tel: +81-466-32-1108. Fax: +81-466-29-4455;
e-mail: mitsunori.kouno@takeda.com
Current affiliation.
b
Axcelead Drug Discovery Partners, Inc., 26-1, Muraoka-Higashi 2-Chome, Fujisawa,
Kanagawa 251-0012, Japan
c Tri-Institutional Therapeutics Discovery Institute, Inc., 413 East 69th Street, New York, NY
10021, U.S.A.

d Japan Tobacco Inc., Central Pharmaceutical Research Institute, 1-1 Murasaki-cho, Takatsuki,
Osaka, 569-1125, Japan.
e
Cardurion Pharmaceuticals K.K. c/o Shonan Research Center, Takeda Pharmaceutical
Company Ltd. 26-1, Muraoka-Higashi 2-chome Fujisawa, Kanagawa 251-8555, Japan
Introduction
The Th17 cells, and the inflammatory cytokines they produce (IL-17A, IL-17F, etc.) play a
central role in the pathogenesis of various autoimmune diseases such as inflammatory bowel
disease, rheumatoid arthritis, multiple sclerosis, and psoriasis.1–5 The biologics targeting IL-17A
and IL-17A receptor have demonstrated clinical efficacy in psoriasis, rheumatoid arthritis, and
uveitis.6–8 Therefore, targeting Th17 cells has emerged as an attractive therapeutic intervention
for these immune-related diseases.
Retinoic acid receptor-related orphan receptor-gamma t (RORγt), which is one of the orphan
nuclear receptors, is a master regulator of Th17, IL-17, and IL-17/IFN-γ co-producing cells
(Th1/17 cells) differentiation.9,10 Expression of the RORγt is thought to be restricted to lymphoid
organs, such as the thymus.11 RORγt contributes to the expression of not only IL-17A, but also
other pro-inflammatory factors such as IL-17F, IL-22, IL-23 receptor, and granulocyte
macrophage colony-stimulating factor, secreted from various immune cells (e.g., Th17, Th1/17,
innate lymphoid cells, and γδ T cells).12,13 Deletion of RORγt results in impaired differentiation of
murine and human Th17 cells. RORγt knockout mice are resistant to various autoimmune
disease models such as colitis and experimental autoimmune encephalomyelitis (EAE).
Therefore, inhibition of RORγt is expected to show treatment effects on various autoimmune
diseases by suppressing differentiation and activation of Th17 and Th1/17 cells.

Recently, a large number of RORγt inhibitors have been reported from many research groups
(Figure 1). It was first reported that natural products such as digoxin (1) and ursolic acid (2)
inhibited RORγt and Th17 cell differentiation as an antagonist, and ameliorated EAE symptoms
in mice by the intraperitoneal administration.14,15 After that, many synthetic small molecular
RORγt inverse agonists have been disclosed in the literatures and patents.16–19 Among them, some
compounds such as VTP-43742, JNJ-3534, ARN-6039, ADZ-0284, and JTE-451 are under
clinical trials.20–23 Examples of the structures of known RORγt inverse agonists (3-7) are shown in
Figure 1.
Previously, we reported a series of phenylglycinamide derivatives as a novel class of ROR t
inverse agonists having a unique binding mode.24 Compound 8a (Figure 1) showed potent in
vitro ROR tt inverse agonistic activity and moderate pharmacokinetic (PK) profile, and
demonstrated inhibitory effect in a mouse IL23-induced IL-17A expression assay at a dose of
100 mg/kg, p.o. For further enhancement of in vivo potency, we embarked the exploration of the
second generation chemotypes with superior in vitro activity and PK profile.
Herein, we report the design and synthesis of novel tetrahydroisoquinoline derivatives as
potent and orally available ROR t inverse agonists obtained from the cyclization of the central
phenylglycinamide core of compound 8a by structure-based design, and subsequent optimization
of right-hand acyl moiety. Furthermore, the biological activities and in vivo pharmacodynamic
(PD) efficacy of compound 15 in mice are described.

Figure 1. Chemical structures of reported ROR t inhibitors
Result and Discussion
Design Strategy Starting with Phenylglycine Derivative. For further enhancement of in vivo
potency of the phenylglycinamide series, we initially focused on the flexible central diamide
structure and hypothesized that reducing rotatable bond to rigidify the molecule would improve
in vitro potency and PK profile.25 In the course of optimizing phenylglycinamide derivatives, we
succeeded in determining the crystal structure of ROR t protein in complex with
phenylglycinamide analogue 8b. According to the crystal structure24, it was indicated that the
central phenyl ring could be connected with the N-methyl amide moiety by some linkers without
affecting the tridental conformation of the entire molecule including the key interactions with
near amino acid residues (Figure 2). Hence, we focused on cyclization between the central
phenyl ring and N-methyl amide moiety, and designed tetrahydroisoquinoline (Y) and the 1,4-
benzoxazepine (Z) scaffold as novel bicyclic compounds which enable to reduce free rotatable
bonds and thereby rigidify the structure (Figure 3).

Figure 2. Surface view of the crystal structure of ROR t protein in complex with
phenylglycinamide analogue 8b.
Figure 3. Cyclization strategy starting from the phenylglycinamide core and designed scaffolds
Initial SAR and Design Concept Validation. The synthesized compounds were initially
evaluated using a RORγt biochemical binding assay and a RORγt jurkat cellular reporter gene
assay. In the binding assay (cell-free system), our original ligand with BODIPY label was used

for evaluating the affinity of the test compounds for human RORγt by time-resolved
fluorescence resonance energy transfer (TR-FRET) technology. In the reporter gene assay (cell-
based), human Jurkat cells were used for evaluating the functional ability of the test compounds
to affect transcriptional activity of RORγt by measuring luciferase activity. The PK profiles of
the compounds were evaluated in mouse cassette-dosing tests. The in vivo PD efficacy of
selected compounds was assessed by the action on IL-23-induced gene expression in mice.
First, we examined the cyclization of phenylglycinamide derivative 8a into six-membered
tetrahydroisoquinoline derivative 9 and seven-membered 1,4-benzoxazepine derivative 10. The
results are summarized in Table 1. As we expected, cyclization to the bicyclic core from
phenylglycinamide did not affect the biochemical and cellular potencies, and the compound 9
showed slightly improved cellular reporter activity and equipotent binding activity to compound
8a. In addition to the [6,6]-fused ring, the [6,7]-fused 1,4-benzoxazepine derivative 10 also
showed tolerable biochemical binding and cellular reporter activity. However, conversion to
compound 10 resulted in a much poorer PK profile compared with phenylglycinamide 8a.
Along with the central core modification, we also conducted the transformation of the
hydroxyisoxazole moiety. In our previous efforts, we found that the hydroxyisoxazole group was
suitable for the acyl moiety in the phenylglycinamide series. The 3-hydroxyisoxazole group is a
commonly known carboxylic acid bioisostere, and widely used in medicinal chemistry drug
design.26 Thus, we conducted introduction of a carboxylic acid group by bioisosteric
transformation of the acyl moiety. Although the carboxylic acid derivative 12 had slightly
decreased reporter activity in spite of potent binding, the compound showed remarkable
improvement of plasma exposure and oral availability. It was of interest that there is no
significant difference in the PK profile between hydroxyisoxazole and carboxylic acid
derivatives of phenylglycinamide compounds (8a vs 11). It was also found that the ring closure

of the phenylglycinamide 8a to form the tetrahydroisoquinoline did not increase plasma exposure
(9). These results suggested that the combination of the carboxylic acid moiety and central fused
bicyclic ring is very important for the PK profile. Thus, we successfully conducted scaffold
exchange of phenylglycinamide derivatives and generated a new [6,6]-fused ring core scaffold of
ROR t inverse agonist. We decided to focus on the tetrahydroisoquinoline carboxylic acid
derivatives and conducted further optimization on the acyl tether.
Table 1. In vitro activities and their PK profiles of derivatives modifying core scaffold and acyl
tether

Optimization of carboxylic acid tether moiety. The SAR trends from the study on the acyl
tether are described in Table 2. Initially, we attempted to fix the conformation of the tether and
designed α,β-unsaturated carboxylic acids (13 and 14). The trans-isomer 14 possessed
comparable binding and reporter activity to compound 12, whereas the cis-isomer 13 resulted in
slight loss of binding and reporter activity. However, plasma exposure of compound 14 was not
as large as that of compound 12. Next, we investigated the linker length (compound 12 possesses
two atoms between carbonyl and carboxylate). The three-atom-linker analog 15 and the four-
atom-linker analog 16 showed equipotent activity to 12. Interestingly, one carbon elongation had

a dramatic impact on PK profile, and the compound 15 demonstrated high plasma exposure and
excellent bioavailability. Replacement of the carboxylic acid with a carbamoyl group (17) led to
an increase of biochemical potency, and 17 showed the most potent reporter activity in this series.
However, the non-acidic substituent resulted in a poor PK profile. On the basis of these results,
we focused on the three atom linker and performed linker modification. Regarding the
introduction of a methyl group, the monomethyl analog 18 retained potent binding and reporter
activity and good PK profile whereas the gem-dimethyl analog 19 showed decreased reporter
activity. Replacement of the methyl group of 19 with a hydroxy group restored reporter activity,
but the plasma exposure was decreased (20). In addition to substituent introduction, we changed
the linker from carbon to oxygen or nitrogen. However, these modifications resulted in decrease
of reporter activity (21 and 22). As for further bioisosteric transformation, thiazolidinedione
instead of carboxylic acid showed potent activities, but the PK profile got worse (23). From these
results, we concluded that compound 15 was the best compound in the tetrahydroisoquinoline
series, and we selected 15 for further in vivo biological evaluation.
Table 2. In vitro activities and their PK profiles of derivatives modifying the acyl tether

O
O
H
F
N
N
O
Si
R
F
Bindinga
Reporter geneb
mouse PKd
compound
R
AUC0-8h
(ng·h/mL)
F
(%)
IC50 (nM)
[95%CI]c
IC50 (nM)
[95%CI]c
4.2
[3.6-4.8]
54
[41-70]
OH
2500
51
N.T.
51
12
13
14
O
O
17
[10-27]
320
[260-410]
N.T.
OH
OH
4.1
[3.4-4.9]
38
[29-49]
619
O
O
3.0
[2.4-3.8]
23
[19-29]
6894
1031
94
84
15
16
OH
OH
6.6
[5.6-7.8]
24
[19-31]
O
O
O
3.1
[2.3-4.0]
6.7
[4.7-9.6]
17
18
92
14
65
NH2
3.8
[3.1-4.7]
19
[12-30]
2498
OH
O
O
5.6
[4.9-6.5]
46
[28-78]
2273
159
40
34
19
20
21
OH
OH
HO
4.5
[3.8-5.3]
32
[25-41]
O
O
3.5
[3.0-4.0]
140
[97-190]
N.T.
N.T.
N.T.
N.T.
O
N
OH
OH
7.8
[6.5-9.4]
220
[150-320]
22
23
O
7.6
[6.4-9.2]
41
[28-62]
S
120
13
NH
O
a Displacement assay of BODIPY-labeled ligand with apo ROR t b Inhibition of ROR t transcriptional activity
human Jurkat cells c 95% confidence intervals (CI) conducted in duplicate (n = 2) d The values shown as means
of three determinations (1 mg/kg, po)

In vivo PD Study and PK Profile of Compound 15. To determine whether a ROR t inverse
agonist could modulate the cytokine production through suppressing differentiation and
activation of Th17 and Th1/17 cells, an in vivo mouse PD model was developed. We established
Mea mouse IL-23-induced cytokine expression model.27 This PD model was used to evaluate the
effect of the compounds on IL-23-induced IL-17A expression in the ears of mice. The results are
shown in Figure 5. Compound 15 showed robust and dose-dependent inhibition of IL-17A
expression after oral administration at 3, 10, and 30 mg/kg (ED50= 2.8 mg/kg). On the other hand,
we also investigated the expression of IFN- which is not regulated by ROR t as for counter
assay, and compound 15 did not affect the expression of IFN- (data not shown). Moreover, the
tetrahydroisoquinoline series represented a significant improvement of in vivo PD efficacy
relative to phenylglycinamide derivative 8a, which showed PD efficacy only at a dose of 100
mg/kg, p.o.. In parallel with the PD assay, we measured the plasma concentration of compound
15 after oral administration. Compound 15 demonstrated excellent PK profile in mice, high
maximum plasma concentration (Cmax: 1.63, 7.72, 19.0 µg/mL), and plasma exposure (AUC0-24h
:
10.7, 48.0, 159 µg·h/mL) increased in a dose-dependent manner (at dose of 3, 10, 30 mg/kg, p.o.).

160
140
120
100
80
#
60
40
*
20
*
0
N
V
3
10
30
Compound 15 (mg/kg)
Figure 5. Assessment of 15 in mice IL-23-induced cytokine expression PD assay. Data are
represented as the mean values s.e. of 7 separate experiments. N indicates the PBS-injected and
V indicates the IL-23-injected and vehicle-administered control group. #p<0.05 (Aspin-Welch
test) compared with PBS-treated group (N) *p<0.025 (one-tailed Shirley Williams' test)
compared to the IL-23-injected and vehicle-administered group (V).

Chemistry.
Synthesis of the tetrahydroisoquinoline-aniline unit28 is described in Scheme 1. 3-
Methoxyphenethylamine 24 was reacted with glyoxylic acid ethyl ester in the presence of HBr to
afford tetrahydroisoquinoline ethyl ester 25. Boc protection and O-methylation proceeded
smoothly, and subsequent hydrolysis by using LiOH gave the carboxylic acid 28. Amidation
with a poor nucleophilic 3,5-difluoro-4-trimethylsilylaniline was achieved by using
propylphosphonic acid anhydride cyclic trimer (T3P) in the presence of a stoichiometric amount
of DMAP to obtain racemic intermediate 29.
Scheme 1a
aReagents and conditions: (a) glyoxilic acid ethyl ester, HBr, EtOH, reflux, 70%; (b) Boc2O,
Et3N, THF, H2O, rt, 87%; (c) MeI, Cs2CO3, DMF, rt, 96%; (d) LiOH, THF, EtOH, rt, 94%; (e)
3,5-difluoro-4-trimethylsilyl aniline, T3P, DMAP, EtOAc, 80 °C, 60%
Synthesis of target compounds 15 and 34, and the determination of absolute configuration for
the tetrahydroisoquinoline-aniline unit were illustrated in Scheme 2. First, we conducted chiral
HPLC resolution of the racemate 29 and prepared the optically pure compounds 30 and 32.
Deprotection of amino group was carefully conducted in short time due to the instability of
trimethylsilyl group under acidic conditions, and the chiral amines 31 and 33 were obtained. The
target compounds (15 and 34) were synthesized from compounds 31 and 33 by the reaction with
glutaric acid anhydride. The absolute configuration of 31 was determined as (R)-configuration by
X-ray crystallographic analysis of compound 31 tartrate (Figure 5). Therefore, the eutomer 15
was proved to be (R)-configuration.

Scheme 2a
aReagents and conditions: (a) prep. HPLC, CHIRALPAK AD, hexane/EtOH, 42% for 30 and
44% for 32; (b) TFA, rt, 93% for 31 and 91% for 33; (c) glutaric anhydride, Et3N, THF, 56% for
15 and 49% for 34
Figure 6. ORTEP of tartrate of compound 31, thermal ellipsoids are drawn at 50% probability.

Synthesis of the target compounds is described in Scheme 3. The final compounds 9, 11-16,
and 18-23 were afforded via acylation with the corresponding carboxylic acids or reaction with
the corresponding acid anhydrides. Carboxamide derivative 17 was obtained by amidation from
15 using HATU and ammonium chloride.
Scheme 3a
1
i
aReagents and conditions: (a) R -CO2H, HATU, Pr2NEt, DMF, 12–52% for 9, 13, 14, 23; (b)
acid anhydrides, Et3N, THF, 29–95% for 12, 15, 16, 18-22; (c) ammonium chloride, HATU,
iPr2NEt, DMF, 57%
Preparation of the benzoxazepine derivative 10 is illustrated in Scheme 4. Boc protection of
N-benzylethanolamine 36 gave the compound 37. The Mitsunobu reaction with 3-
methoxyphenol using DIAD and PPh3, and subsequent deprotection provided the intermediate 39.
In situ generation of methylbromo(methoxy)acetate followed by the reaction with 39 afforded a
coupling product 40. The benzoxazepine scaffold was then constructed by TMSCl-promoted
intramolecular cyclization29 at room temperature to yield compound 41. The benzyl group was
removed by hydrogenation using Pd(OH)2-C, followed by protection with a Boc group and
hydrolysis to give 43. Amidation under the same conditions as synthesis of
tetrahydroisoquinoline derivative afforded amide 44. The racemate of 10 was synthesized by

acidic deprotection of 44 and subsequent acylation with 3-hydroxyisoxazole-5-carboxylic acid.
The target eutomer was obtained by chiral HPLC resolution and gave the optically pure
compound 10.
Scheme 4a
aReagents and conditions: (a) (Boc)2O, THF, rt, quant.; (b) 3-methoxyphenol, DIAD, PPh3, THF,
i
0 °C–rt, 41%; (c) TFA, 0 °C–rt, quant.; (d) methyl methoxyacetate, NBS, AIBN/PhCF3, Pr2NEt,
THF, rt, 76%; (e) TMSCl, MeCN, 0 °C–rt, 60%; (f) H2, Pd(OH)2-C, MeOH, rt, 96%; (g)
(Boc)2O, THF, rt, quant. 77% from 42; (h) LiOH, MeOH-THF, rt, 94%; (i) 3,5-difluoro-4-
trimethylsilyl aniline, T3P, DMAP, EtOAc, 80 °C, 81%; (j) TFA, rt, 88%; (k) 3-
i
hydroxyisoxazole-5-carboxylic acid, HATU, Pr2NEt, DMF, 41%; (l) prep. SFC, CHIRALPAK
AS-H, CO2/EtOH, 30%
Conclusion
In the course of our exploration of novel RORγt inverse agonists that are therapeutically
effective against various autoimmune diseases, we designed, synthesized, and evaluated a series
of tetrahydroisoquinoline derivatives. Cyclization of the phenylglycinamide core by SBDD

approach successfully delivered a novel bicyclic scaffold with tolerable biochemical binding and
cellular reporter activity. Combination of a carboxylic acid tether and a tetrahydroisoquinoline
scaffold resulted in significant improvement of the PK profile. The optimization of the acyl
tether led to the discovery of 5-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-
methoxy-3,4-dihydroisoquinolin-2(1H)-yl)-5-oxopentanoic acid (15), a potent, and orally active
RORγt inverse agonist with excellent PK profile. Oral administration of compound 15 to mice
robustly and dose-dependently inhibited IL-17A production in an IL23-induced gene expression
assay. The results suggested that our potent and selective RORγt inverse agonists would become
a beneficial agent for the treatment of Th17-driven autoimmune diseases.

Experimental section
Chemistry
Reagents and solvents were obtained from commercial sources and used without further
purification. Reaction progress was determined by thin layer chromatography (TLC) analysis on
Merck Kieselgel 60 F254 plates or Fuji Silysia NH plates. Silica gel column chromatography
was performed on Purif-Pack (SI or NH, SHOKO SCIENTIFIC). Proton nuclear magnetic
resonance (1H NMR) spectra were recorded on Bruker Ultra Shield-300 (300 MHz) instruments.
Chemical shifts are given in parts per million (ppm) with tetramethylsilane as an internal
standard. Abbreviations are used as follows: s = singlet, d = doublet, t = triplet, q = quartet, m =
multiplet, dd = doublets of doublet, br = broad. Coupling constants (J values) are given in hertz
(Hz). LC–MS analysis was performed on a Shimadzu LC-20AD separations module or Agilent
1200 series, operating in ESI (+ or -) ionization mode. Analytes were eluted using a linear
gradient of 0.05% TFA containing H2O/CH3CN or 0.01% TFA containing H2O/CH3CN or 5 mM
ammonium acetate containing H2O/CH3CN or 10 mM NH4HCO3 containing H2O/CH3CN or
0.01% heptafluorobutyric acid /1.0% isopropyl alcohol containing H2O/CH3CN mobile phase.
Preparative HPLC was performed on a Waters 2525 separations module (L-column2 ODS (20
x150 mm I.D., CERI, Japan); MS spectra were recorded using a Waters ZQ2000 with
electrospray ionization or on a GILSON system, equipped with a L-column2 ODS (20 x150 mm
I.D., CERI, Japan) or on Waters Deltaprep 300 system or on Shimadzu 10A VP system. Samples
were eluted using a linear gradient of 0.1% TFA in H2O/CH3CN or 10 mM NH4HCO3 in H2O
/CH3CN, or H2O/CH3CN. Purity data were collected by a HPLC with Corona CAD (Charged
Aerosol Detector), Nano quantity analyte detector (NQAD), or photo diode array detector. The
column was a Capcell Pak C18AQ (50 mm x 3.0 mm I.D., Shiseido, Japan) or L-column 2 ODS
(30 mm x 2.0 mm I.D., CERI, Japan) with a temperature of 50 °C and a flow rate of 0.5 mL/min.

Mobile phase A and B under a neutral condition were a mixture of 50 mM Ammonium acetate,
H2O and CH3CN (1:8:1, v/v/v) and a mixture of 50 mM ammonium acetate and CH3CN (1:9, v/v),
respectively. The ratio of mobile phase B was increased linearly from 5% to 95% over 3 min,
95% over the next 1 min. Mobile phase A and B under an acidic condition were a mixture of
0.2% formic acid in 10 mM ammonium formate and 0.2% formic acid in CH3CN, respectively.
The ratio of mobile phase B was increased linearly from 14% to 86% over 3 min, 86% over the
next 1 min.
Ethyl 6-hydroxy-1,2,3,4-tetrahydroisoquinoline-1-carboxylate hydrobromide (25). 48 %
Hydrobromic acid (101 g, 600 mmol) was added dropwise (over a period of 10 min) to m-
methoxyphenethylamine (22.68 g, 150 mmol) at room temperature. After being stirred at 120 °C
for 12 h, the reaction mixture was diluted with EtOH (200 ml) and concentrated in vacuo. The
evaporation was repeated after the addition of toluene (200 ml, 4 times) to give white solid. The
residue was dissolved in EtOH (150 ml), and 47 % glyoxylic acid ethyl ester (polymer form)
(35.8 g, 165 mmol) was added thereto. After being refluxed for 15 h, the reaction mixture was
concentrated in vacuo. The evaporation was repeated after the addition of toluene (200 ml, twice).
The residual oil was diluted with EtOAc (200 ml) and sitrred at room temperature for 40 min to
give crystals, which were collected by filtration and washed with EtOAc to give 25 (31.8 g, 70 %)
1
as a pale yellow solid. H NMR (300 MHz, DMSO-d6) 1.26 (3H, t, J = 7.2 Hz), 2.86 – 2.99
(2H, m), 3.35 – 3.61 (4H, m), 4.19 – 4.35 (2H, m), 5.35 (1H, s), 6.63 (1H, d, J = 2.6 Hz), 6.73
(1H, dd, J = 8.7, 2.6 Hz), 7.23 (1H, d, J = 8.3 Hz), 9.40 (1H, brs), 9.69 (2H, brs). LC/MS m/z
222.2 (M + H).
2-tert-Butyl 1-ethyl 6-hydoxy-3,4-dihydroisoquinoline-1,2(1H)-dicarboxylate (26). To a
solution of 25 (31.7 g, 105 mmol) in THF (300 ml) and water (80 ml) was added Et3N (14.6 ml,

105 mmol) and Boc2O (25.6 ml, 110 mmol) at room temperature. The mixture was stirred at
room temperaturefor 1 h. The mixture was poured into water and extracted with EtOAc. The
organic layer was separated, washed with brine, dried over MgSO4 and concentrated in vacuo.
The residue was purified by column chromatography (silica gel, eluted with 0%–50% EtOAc in
1
hexane) to give 26 (29.5 g, 87 %) as a colorless oil. H NMR (300 MHz, CDCl3) 1.17 – 1.32
(4H, m), 1.45 – 1.53 (10H, m), 2.69 – 2.99 (2H, m), 3.62 – 3.83 (2H, m), 4.07 – 4.20 (2H, m),
4.93 – 5.03 (1H, m), 5.33 (1H, s), 6.63 (1H, s), 6.69 (1H, d, J = 8.7 Hz), 7.31 – 7.39 (1H, m).
LC/MS m/z 222.1 (M + H – (Boc)).
2-tert-Butyl 1-ethyl 6-methoxy-3,4-dihydroisoquinoline-1,2(1H)-dicarboxylate (27). To a
solution of 26 (6.43 g, 20 mmol) in DMF (50 ml) was added Cs2CO3 (7.82 g, 24.0 mmol) and
MeI (2.50 ml, 40.0 mmol) at room temperature. The mixture was stirred at room temperature
overnight. The mixture was poured into water and extracted with EtOAc. The organic layer was
separated, washed with water and brine, dried over MgSO4 and concentrated in vacuo. The
residue was purified by column chromatography (NH silica gel, eluted with 0%–20% EtOAc in
1
hexane) to give 27 (6.45 g, 96 %) as a colorless oil. H NMR (300 MHz, CDCl3) 1.16 – 1.31
(3H, m), 1.40 – 1.52 (9H, m), 2.71 – 3.02 (2H, m), 3.64 – 3.84 (5H, m), 4.07 – 4.20 (2H, m),
5.34 (1H, s), 6.68 (1H, s), 6.77 (1H, dd, J = 8.3, 2.6 Hz), 7.35 – 7.45 (1H, m).
2-(tert-Butoxycarbonyl)-6-methoxy-1,2,3,4-tetrahydroisoquinoline-1-carboxylic acid (28).
To a solution of 27 (6.37 g, 19 mmol) in THF (20 ml) and EtOH (20 ml) was added NaOH (19.0
ml, 38.0 mmol) at room temperature. The mixture was stirred at room temperature overnight.
After evaporation of the solvent, the residue was neutralized with 2 N HCl at 0 °C and extracted
with EtOAc. The organic layer was separated, washed with water and brine, dried over MgSO4
1
and concentrated in vacuo to give 28 (5.50 g, 94 %) as a white amorphous solid. H NMR (300

MHz, CDCl3) 1.48 (3H, s), 1.44 (6H, s), 2.79 (1H, d, J = 15.9 Hz), 2.93 (1H, d, J = 15.9 Hz),
3.55 – 3.68 (1H, m), 3.68 – 3.91 (4H, m), 5.34 (1H, s), 6.68 (1H, d, J = 2.6 Hz), 6.77 (1H, dd, J =
8.5, 2.5 Hz), 7.38 (1H, d, J = 8.7 Hz). LC/MS m/z 208.1 (M + H – (Boc)).
tert-Butyl
1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
To mixture of 3,5-difluoro-4-
dihydroisoquinoline-2(1H)-carboxylate (29).
a
(trimethylsilyl)aniline (1.05 g, 5.22 mmol), 28 (1.76 g, 5.74 mmol), DMAP (0.70 g, 5.74 mmol),
and N,N'-diisopropylethylamine (4.56 ml, 26.1 mmol) in EtOAc (30 ml) was added T3P, 1.7 M
solution in EtOAc (9.21 ml, 15.7 mmol). After being stirred at 80 °C for 2 h, the mixture was
quenched with water at ambient temperature and extracted with EtOAc. The organic layer was
separated, washed with 10 % aqueous citric acid, dried over MgSO4 and concentrated in vacuo.
1
The residue was triturated with Et2O to give 29 (1.41 g, 55 %) as a white powder. H NMR (300
MHz, CDCl3) 1.52 (9H, s), 2.76 – 2.98 (2H, m), 3.51 – 3.75 (2H, m), 3.80 (3H, s), 5.58 (1H,
brs), 6.73 (1H, d, J = 2.3 Hz), 6.81 (1H, dd, J = 8.3, 2.3 Hz), 6.97 – 7.07 (2H, m), 7.19 (1H, brs),
9.11 (1H, brs). LC/MS m/z 489.3 (M + H)
tert-butyl
(1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinoline-2(1H)-carboxylate (30). Compound 29 (1.44 g) was subjected to
preparative chiral HPLC (Column: CHIRALPAK AD, Eluent: Hexane/ethanol =850/150 (v/v),
Flow rate: 60 mL/min, Pressure: 0.2 Mpa, Detector&sens.: UV220 nM, Temperature: 30 °C).
The factions with a shorter retention time gave the title compound 30 (610 mg, 42%) with >
99%ee as colorless amorphous solid. LC/MS m/z 489.3 (M + H).
(1R)-N-(3,5-difluoro-4-(trimethylsilyl)phenyl)-6-methoxy-1,2,3,4-tetrahydroisoquinoline-1-
carboxamide (31). Ice-cooled trifluoroacetic acid (5.0 ml, 1.24 mmol) was added to 30 (610 mg,

1.24 mmol) at room temperature. After being stirred at room temperature for 2 min, the reaction
mixture was poured into iced saturated aqueous NaHCO3 and extracted with EtOAc. The
organic layer was dried over MgSO4 and concentrated in vacuo to give 31 (450 mg, 93 %) as a
colorless amorphous solid. This product was subjected to the next reaction without further
purification. LC/MS m/z 389.2 (M – H)
5-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-5-oxopentanoic acid (15). A mixture of 31 (300 mg, 0.77
mmol), dihydro-2H-pyran-2,6(3H)-dione (131 mg, 1.15 mmol), TEA (0.214 ml, 1.54 mmol), and
THF (6.0 ml) was stirrred at room temperature for 3 h. The mixture was concentrated in vacuo.
The residue was purified by column chromatography (diol column, eluted with 50%–100%
1
EtOAc in hexane) to give 15 (217 mg, 56 %) as a colorless amorphous solid. H NMR (300
MHz, DMSO-d6) 0.30 (9H, s), 1.64 – 1.85 (2H, m), 2.23 – 2.33 (2H, m), 2.36 – 2.62 (2H, m),
2.73 – 2.88 (1H, m), 3.05 –.19 (1H, m), 3.44 – 3.57 (1H, m), 3.73 (3H, s), 3.94 – 4.08 (1H, m),
5.58 – 5.64 (1H, m), 6.77 – 6.86 (2H, m), 7.15 – 7.25 (2H, m), 7.46 (1H, d, J = 9.1 Hz), 10.76
(1H, s), 12.03 (1H, s). LC/MS m/z 505.4 (M + H). Optical rotation: [α]25 –6.6 (c 0.250,
D
MeOH). HPLC purity: 98.9%.
tert-butyl
(1S)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinoline-2(1H)-carboxylate (32). Compound 29 (1.44 g) was subjected to
preparative chiral HPLC (Column: CHIRALPAK AD, Eluent: Hexane/ethanol =850/150 (v/v),
Flow rate: 60 mL/min, Pressure: 0.2 Mpa, Detector&sens.: UV220 nM, Temperature: 30 °C).
The factions with a longer retention time gave the title compound 32 (630 mg, 44%) with >
99%ee as colorless amorphous solid. LC/MS m/z 489.3 (M + H).

(1S)-N-(3,5-difluoro-4-(trimethylsilyl)phenyl)-6-methoxy-1,2,3,4-tetrahydroisoquinoline-1-
carboxamide (33). Compound 33 was prepared in a manner similar to that described for 31 in
91% yield as a colorless amorphous solid. LC/MS m/z 389.2 (M – H)
5-((1S)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-5-oxopentanoic acid (34). Compound 34 was prepared in a
1
manner similar to that described for 15 in 49% yield as a colorless amorphous solid. H NMR
(300 MHz, DMSO-d6) 0.29 (9H, s), 1.63 – 1.82 (2H, m), 2.18 – 2.32 (3H, m), 2.36 – 2.49 (1H,
m), 2.52 – 2.61 (1H, m), 2.72 – 2.88 (1H, m), 3.05 – 3.19 (1H, m), 3.43 – 3.56 (1H, m), 3.73
(3H, s), 5.56 – 5.67 (1H, m), 6.77 – 6.86 (2H, m), 7.15 – 7.25 (2H, m), 7.46 (1H, d, J = 9.1 Hz),
10.76 (1H, s), 12.01 (1H, brs). LC/MS m/z 503.3 (M – H) Optical rotation: [α]25 +7.2 (c
D
0.252, MeOH). HPLC purity: 90.5%.
(1R)-N-(3,5-difluoro-4-(trimethylsilyl)phenyl)-2-((3-hydroxy-1,2-oxazol-5-yl)carbonyl)-6-
methoxy-1,2,3,4-tetrahydroisoquinoline-1-carboxamide (9). To a stirred solution of 31 (158
mg, 0.40 mmol), N,N'-diisopropylethylamine (0.141 ml, 0.81 mmol), and 3-hydroxyisoxazole-
5-carboxylic acid (54.8 mg, 0.42 mmol) in DMF (3.0 ml) was added HATU (185 mg, 0.49
mmol) at room temperature. The mixture was stirred at room temperature overnight, and the
mixture was quenched with water and extracted with EtOAc. The organic layer was separated,
washed with water and brine, dried over MgSO4 and concentrated in vacuo. The residue was
purified by column chromatography (silica gel, eluted with 20%–90%EtOAc in hexane) and
crystallized from EtOAc–hexane to give 9 (53.5 mg, 26 %) as a colorless amorphous solid. 1H
NMR (300 MHz, DMSO-d6) 0.30 (9H, s), 2.79 – 2.96 (1H, m), 3.06 – 3.22 (1H, m), 3.74 (4H,
s), 4.06 – 4.24 (1H, m), 5.65 (1H, s), 6.55 (1H, s), 6.80 – 6.92 (2H, m), 7.11 – 7.26 (2H, m),

7.51 (1H, d, J = 9.4 Hz), 10.92 (1H, s), 11.97 (1H, s). LC/MS m/z 500.1 (M – H). Optical
rotation: [α]25D –15.2 (c 0.128, MeOH). HPLC purity: 91.4%.
4-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-4-oxobutanoic acid (12). Compound 12 was prepared in a
manner similar to that described for 15, using dihydrofuran-2,5-dione in 95% yield as a colorless
amorphous solid. 1H NMR (300 MHz, DMSO-d6) 0.29 (9H, s), 2.57 – 2.92 (6H, m), 3.03 –
3.23 (1H, m), 3.45 – 3.63 (1H, m), 3.73 (3H, s), 3.97 – 4.10 (1H, m), 5.56 – 5.70 (1H, m), 6.76 –
6.89 (2H, m), 7.13 – 7.28 (2H, m), 7.40 – 7.50 (1H, m), 10.63 – 10.87 (1H, m). LC/MS m/z
489.2 (M – H). Optical rotation: [α]25D +3.8 (c 0.252, MeOH). HPLC purity: 94.8%.
(2Z)-4-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-4-oxobut-2-enoic acid (13). Compound 13 was prepared in a
manner similar to that described for 9, using maleic acid in 12% yield as a colorless amorphous
1
solid. H NMR (300 MHz, DMSO-d6) 0.30 (9H, s), 2.68 – 2.89 (1H, m), 2.92 – 3.08 (1H, m),
3.40 – 3.52 (1H, m), 3.73 (3H, s), 3.85 – 4.05 (1H, m), 5.70 (1H, s), 6.07 (1H, d, J = 12.1 Hz),
6.75 – 6.87 (3H, m), 7.20 – 7.31 (2H, m), 7.43 (1H, d, J = 8.3 Hz), 10.65 – 10.81 (1H, m), 12.82
(1H, brs). LC/MS m/z 487.2 (M – H). Optical rotation: [α]25D +73.0 (c 0.127, MeOH). HPLC
purity: 96.9%.
(2E)-4-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-4-oxobut-2-enoic acid (14). Compound 14 was prepared in a
manner similar to that described for 9, using fumaric acid in 18% yield as a colorless amorphous
1
solid. H NMR (300 MHz, DMSO-d6) 0.30 (9H, s), 2.78 – 2.93 (1H, m), 3.06 – 3.21 (1H, m),
3.60 – 3.71 (1H, m), 3.73 (3H, s), 4.08 – 4.21 (1H, m), 5.63 – 5.79 (1H, m), 6.51 – 6.62 (1H, m),

6.77 – 6.90 (2H, m), 7.13 – 7.25 (2H, m), 7.39 – 7.57 (2H, m), 10.84 (1H, s), 13.07 (1H, brs).
LC/MS m/z 487.2 (M – H). Optical rotation: [α]25 +7.7 (c 0.127, MeOH). HPLC purity:
D
99.3%.
6-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-6-oxohexanoic acid (16). Compound 16 was prepared in a
manner similar to that described for 15, using oxepane-2,7-dione in 11% yield as a colorless
amorphous solid. 1H NMR (300 MHz, DMSO-d6) 0.29 (9H, s), 1.41 – 1.67 (4H, m), 2.12 –
2.31 (1H, m), 2.36 – 2.61 (3H, m), 2.66 – 2.89 (1H, m), 3.00 – 3.19 (1H, m), 3.43 – 3.60 (1H,
m), 3.72 (3H, s), 3.95 – 4.11 (1H, m), 5.61 (1H, s), 6.82 (2H, d, J = 9.8 Hz), 7.19 (2H, d, J = 9.8
Hz), 7.45 (1H, d, J = 8.3 Hz), 10.74 (1H, s), 11.97 (1H, s). LC/MS m/z 517.3 (M – H). Optical
rotation: [α]25D –3.3 (c 0.253, MeOH). HPLC purity: 96.7%.
(1R)-2-(5-amino-5-oxopentanoyl)-N-(3,5-difluoro-4-(trimethylsilyl)phenyl)-6-methoxy-
1,2,3,4-tetrahydroisoquinoline-1-carboxamide (17). To a stirred solution of 15 (200 mg, 0.40
mmol) and N,N'-diisopropylethylamine (0.20 ml, 1.19 mmol) in DMF (4.0 ml) was added
ammonium chloride (25.4 mg, 0.48 mmol) and HATU (181 mg, 0.48 mmol) at room
temperature. The mixture was stirred at room temperature for 3 h. The mixture was quenched
with water and extracted with EtOAc. The organic layer was separated, washed with water and
brine, dried over MgSO4 and concentrated in vacuo. The residue was purified by column
chromatography (silica gel, eluted with 0%–50% EtOAc in hexane) and recrystallized from
1
EtOAc–hexane to give 17 (114 mg, 57 %) as colorless crystals. H NMR (300MHz, DMSO-d6)
0.29 (9H, s), 1.73 (2H, quin, J = 7.1 Hz), 2.05 – 2.16 (2H, m), 2.32 – 2.57 (2H, m), 2.70 – 2.88
(1H, m), 3.04 – 3.18 (1H, m), 3.43 – 3.58 (1H, m), 3.73 (3H, s), 3.94 – 4.08 (1H, m), 5.62 (1H,
s), 6.71 (1H, brs), 6.77 – 6.87 (2H, m), 7.14 – 7.29 (3H, m), 7.46 (1H, d, J = 9.4 Hz), 10.76 (1H,

s). LC/MS m/z 502.3 (M – H). Optical rotation: [α]25D –7.6 (c 0.254, MeOH). HPLC purity:
99.3%.
5-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-3-methyl-5-oxopentanoic acid (18).
Compound 18 was
prepared in a manner similar to that described for 15, using 4-methyldihydro-2H-pyran-2,6(3H)-
1
dione in 46% yield as a colorless amorphous solid. H NMR (300 MHz, DMSO-d6) 0.82 – 0.99
(3H, m), 2.04 – 2.20 (1H, m), 2.21 – 2.41 (3H, m), 2.42 – 2.61 (1H, m), 2.75 – 2.89 (1H, m),
3.03 – 3.21 (1H, m), 3.45 – 3.59 (1H, m), 3.73 (3H, s), 3.97 – 4.10 (1H, m), 5.62 (1H, s), 6.78 –
6.85 (2H, m), 7.14 – 7.25 (2H, m), 7.46 (1H, d, J = 9.4 Hz), 10.77 (1H, s), 12.04 (1H, brs).
LC/MS m/z 517.3 (M – H). Optical rotation: [α]25 +2.8 (c 0.127, MeOH). HPLC purity:
D
99.3%.
5-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-3,3-dimethyl-5-oxopentanoic acid (19). Compound 19 was
prepared in a manner similar to that described for 15, using 4,4-dimethyldihydro-2H-pyran-
1
2,6(3H)-dione in 49% yield as a colorless amorphous solid. H NMR (300 MHz, DMSO-d6)
0.29 (9H, s), 1.09 (6H, d, J = 1.1 Hz), 2.25 – 2.44 (2H, m), 2.56 – 2.86 (2H, m), 3.04 – 3.19 (1H,
m), 3.45 – 3.61 (1H, m), 3.73 (3H, s), 3.96 – 4.15 (2H, m), 5.57 – 5.70 (1H, m), 6.76 – 6.89 (2H,
m), 7.14 – 7.25 (2H, m), 7.45 (1H, d, J = 9.1 Hz), 10.77 (1H, s), 11.93 (1H, brs). LC/MS m/z
531.3 (M – H). Optical rotation: [α]25D +7.3 (c 0.051, MeOH). HPLC purity: 97.5%.
5-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-3-hydroxy-3-methyl-5-oxopentanoic acid (20). Compound 20
was prepared in a manner similar to that described for 15, using 4-hydroxy-4-methyldihydro-2H-

1
pyran-2,6(3H)-dione in 34% yield a pale-yellow amorphous solid. H NMR (300 MHz, DMSO-
d6) 0.29 (9H, s), 1.24 – 1.30 (3H, m), 2.42 – 2.47 (1H, m), 2.71 – 2.87 (6H, m), 3.02 – 3.16 (1H,
m), 3.52 – 3.64 (1H, m), 3.73 (3H, s), 3.96 – 4.16 (1H, m), 5.65 (1H, s), 6.76 – 6.88 (2H, m),
7.16 – 7.28 (2H, m), 7.39 – 7.50 (1H, m), 10.74 (1H, s). LC/MS m/z 535.5 (M + H). Optical
rotation: [α]25D +8.6 (c 0.256, MeOH). HPLC purity: 98.5%.
(2-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-2-oxoethoxy)acetic acid (21). Compound 21 was prepared in a
manner similar to that described for 15, using 1,4-dioxane-2,6-dione in 29% yield a colorless
1
amorphous solid. H NMR (300MHz, DMSO-d6) 0.30 (9H, s), 2.75 – 2.91 (1H, m), 2.99 – 3.17
(1H, m), 3.40 – 3.56 (1H, m), 3.73 (3H, s), 3.91 – 4.02 (1H, m), 4.11 (2H, s), 4.32 – 4.50 (2H,
m), 5.61 (1H, s), 6.75 – 6.89 (2H, m), 7.20 (2H, d, J = 9.8 Hz), 7.40 – 7.50 (1H, m), 10.80 (1H,
s), 12.66 (1H, brs). LC/MS m/z 505.2 (M – H). HPLC purity: 100%.
((2-((1R)-1-((3,5-difluoro-4-(trimethylsilyl)phenyl)carbamoyl)-6-methoxy-3,4-
dihydroisoquinolin-2(1H)-yl)-2-oxoethyl)(methyl)amino)acetic acid (22). Compound 22 was
prepared in a manner similar to that described for 15, using 4-methylmorpholine-2,6-dione in
1
77% yield a colorless amorphous solid. H NMR (300 MHz, DMSO-d6) 0.30 (9H, s), 2.38 (3H,
s), 2.72 – 2.89 (1H, m), 2.89 – 3.00 (1H, m), 3.01 – 3.20 (2H, m), 3.35 – 3.49 (1H, m), 3.51 –
3.66 (3H, m), 3.73 (3H, s), 4.06 – 4.22 (1H, m), 5.60 (1H, s), 6.72 – 6.86 (2H, m), 7.13 – 7.26
(2H, m), 7.39 – 7.50 (1H, m), 10.81 (1H, s). LC/MS m/z 520.4 (M + H). Optical rotation: [α]25
D
+15.5 (c 0.252, MeOH). HPLC purity: 95.2%.
(1R)-N-(3,5-difluoro-4-(trimethylsilyl)phenyl)-2-((2,4-dioxo-1,3-thiazolidin-5-yl)acetyl)-6-
methoxy-1,2,3,4-tetrahydroisoquinoline-1-carboxamide (23). Compound 23 was prepared in

a manner similar to that described for 9, using (2,4-dioxo-1,3-thiazolidin-5-yl)acetic acid in 52%
1
yield as colorless crystals. H NMR (300 MHz, DMSO-d6) 0.29 (9H, s), 2.76 – 2.90 (1H, m),
3.08 – 3.22 (1H, m), 3.24 – 3.38 (2H, m), 3.39 – 3.58 (1H, m), 3.73 (3H, s), 3.94 – 4.09 (1H, m),
4.54 – 4.70 (1H, m), 5.61 (1H, s), 6.77 – 6.90 (2H, m), 7.12 – 7.27 (2H, m), 7.44 (1H, t, J = 8.3
Hz), 10.77 (1H, d, J = 11.7 Hz), 11.96 (1H, brs). LC/MS m/z 546.2 (M – H). Optical rotation:
[α]25D +7.3 (c 0.253, MeOH). HPLC purity: 97.1%.
(R)-4-((2-((3,5-difluoro-4-(trimethylsilyl)phenyl)amino)-1-(4-methoxyphenyl)-2-
oxoethyl)(methyl)amino)-4-oxobutanoic acid (11). Compound 11 was prepared in a manner
similar to that described for 15, using dihydrofuran-2,5-dione in 71% yield as a colorless
amorphous solid. 1H NMR (300 MHz, DMSO-d6) 0.31 (9H, s), 2.43 – 2.49 (2H, m), 2.55 –
2.68 (2H, m), 2.77 (3H, s), 3.75 (3H, s), 6.19 (1H, s), 6.97 (2H, d, J = 8.7 Hz), 7.15 (2H, d, J =
8.7 Hz), 7.24 (2H, d, J = 9.8 Hz), 10.60 (1H, s), 12.07 (1H, brs). LC/MS m/z 477.1 (M – H).
Optical rotation: [α]25D –154.1 (c 0.251, MeOH). HPLC purity: 100%.
tert-Butyl benzyl(2-hydroxyethyl)carbamate (37). Boc2O (16.7 g, 76.4 mmol) was added to a
solution of 2-(benzylamino)ethanol (11.0 g, 72.8 mmol) in THF (220 ml) at room temperature.
After being stirred at room temperature for 16 h, the reaction mixture was concentrated in vacuo.
The residual oil was purified by column chromatography on silica gel (eluted with 10%–60%
1
EtOAc in hexane) to give 37 (18.3 g, quant.) as a colorless oil. H NMR (300 MHz, CDCl3)
0.47 (9H, s), 3.03 (1H, brs), 3.40 (2H, brs), 3.65 – 3.75 (2H, m), 4.48 (2H, brs), 7.21 – 7.37 (5H,
m). LC/MS m/z 152.2 (M + H – (Boc)).
tert-Butyl benzyl(2-(3-methoxyphenoxy)ethyl)carbamate (38). 1.9 M DIAD (47.8 ml, 90.9
mmol) in toluene was added to a solution of 37 (18.3 g, 72.7 mmol), 3-methoxyphenol (7.52 g,