Synthesis and biological evaluation of novel purinyl quinazolinone derivatives as PI3Kδ-specific inhibitors for the treatment of hematologic malignancies

Article abstract of DOI:10.1016/j.bmc.2021.116312

Phosphatidylinositol 3-kinases (PI3Ks) mediate intracellular signal transduction. Aberrant PI3K signaling is associated with oncogenesis and disease progression in solid tumors and hematologic malignancies. Idelalisib (1), a first-in-class PI3Kδ inhibitor for the treatment of hematologic malignancies, was developed, but its sales were limited by black box warnings due to unexpected adverse effects. Therefore, to overcome these adverse events, various quinazolinone derivatives were synthesized and evaluated in vitro based on their inhibitory activity against the PI3K enzyme and the viability of cell lines such as MOLT and SUDHL. Among them, 6f (IC₅₀ = 0.39 nM) and 6m (IC₅₀ = 0.09 nM) showed excellent enzyme activity, and 6m displayed an approximately four-fold higher selectivity for PI3Kγ/δ compared with Idelalisib (1). Furthermore, in vivo PK experiments with 6f and 6m revealed that 6f (AUC_last = 81.04 h*ng/mL, C_max = 18.34 ng/mL, T_max = 0.5 h, t_1/2 = 10.2 h in 1 mpk dose) had improved PK compared with 1. Finally, further experiments will be conducted with 6f selected as a candidate, and the potential for it to be developed as a treatment with good efficacy for hematologic malignancies will be determined.

Full text of DOI:10.1016/j.bmc.2021.116312

Bioorg. Med. Chem. 45 (2021) 116312
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Synthesis and biological evaluation of novel purinyl quinazolinone
derivatives as PI3Kδ-specific inhibitors for the treatment of
hematologic malignancies
,
,
Yeon Su Kima 1, Min Gyeong Cheonb 1, Pulla Reddy Boggua, Su Youn Koha, Gi Min Parka,
Gahee Kimb, Seo Hyun Parkb, Sung Lyea Parkb, Chi Woo Leeb, Jong Woo Kimb
,
,
*
Young Hoon Junga
,
*
a School of Pharmacy, Sungkyunkwan University, Suwon, Gyeonggi-do 16419, Republic of Korea
b Bioway, Inc., Chuncheon, Gangwon-do 24232, Republic of Korea
A R T I C L E I N F O
A B S T R A C T
Keywords:
Phosphatidylinositol 3-kinases (PI3Ks) mediate intracellular signal transduction. Aberrant PI3K signaling is
associated with oncogenesis and disease progression in solid tumors and hematologic malignancies. Idelalisib
(1), a first-in-class PI3Kδ inhibitor for the treatment of hematologic malignancies, was developed, but its sales
were limited by black box warnings due to unexpected adverse effects. Therefore, to overcome these adverse
events, various quinazolinone derivatives were synthesized and evaluated in vitro based on their inhibitory ac-
Quinazolinone
PI3Kδ inhibitor
Hematologic malignancies
tivity against the PI3K enzyme and the viability of cell lines such as MOLT and SUDHL. Among them, 6f (IC50
=
0.39 nM) and 6m (IC50 = 0.09 nM) showed excellent enzyme activity, and 6m displayed an approximately four-
fold higher selectivity for PI3Kγ/δ compared with Idelalisib (1). Furthermore, in vivo PK experiments with 6f and
6m revealed that 6f (AUClast = 81.04 h*ng/mL, Cmax = 18.34 ng/mL, Tmax = 0.5 h, t1/2 = 10.2 h in 1 mpk dose)
had improved PK compared with 1. Finally, further experiments will be conducted with 6f selected as a
candidate, and the potential for it to be developed as a treatment with good efficacy for hematologic malig-
nancies will be determined.
1. Introduction
progress.24–26
Based on the catalytic and regulatory subunits, the class I PI3Ks
Phosphatidylinositol 3-kinases (PI3Ks) are lipid kinases that catalyze
the conversion of phosphatidylinositol 4,5-bisphosphate (PIP2) to
phosphatidylinositol 3,4,5-trisphosphate (PIP3) by phosphorylation and
have been associated with an intracellular signaling pathway, including
cell growth, proliferation, motility, survival, and intracellular trafficking
1–4. The PI3K pathway is dysregulated in various cancers, and aberrant
PI3K signaling is associated with oncogenesis and disease progression
family is divided into two categories.20,27 Class IA PI3Ks are mainly
activated by receptor tyrosine kinases (RTKs), which are heterodimeric
enzymes and composed of an 85 kD regulatory subunit (p85) and a 110
kD catalytic subunit (p110
α
, p110β, p110δ28), whereas class IB PI3Ks,
which are mainly activated by G-protein coupled receptors (GPCRs),
include two different regulatory subunits (p101 or p87) and a single
catalytic subunit (p110γ).29 Among the catalytic subunits, p110
α
involving solid tumors and hematologic malignancies 5–10. Therefore,
(PI3Kα) and p110β (PI3Kβ) are ubiquitous, whereas p110δ (PI3Kδ) and
11–14
PI3K inhibitors
are currently under investigation as potential
p110γ (PI3Kγ) are only expressed in leukocytes, neuroblastoma cells and
hematopoietic cells, respectively. In particular, PI3Kδ regulates the
signaling pathway to induce pathogenesis of CLL, rheumatoid arthritis,
and allergic asthma mediated via B and T cells.30–33
15–16
cancer treatments. For example, pan-PI3K inhibitors
suppress all
class I PI3K isoforms compared with isoform-selective PI3K inhibitors
and dual PI3K/mTOR inhibitors (Fig. 1) 23. In addition, various
17–22
PI3K inhibitors are still being studied and numerous clinical trials are in
Idelalisib (1),34 a first-in-class PI3Kδ inhibitor for the treatment of
* Corresponding authors.
E-mail addresses: jwkim@bioway.co.kr (J.W. Kim), yhjung@skku.edu (Y.H. Jung).
1
These authors equally contributed to this work.
https://doi.org/10.1016/j.bmc.2021.116312
Received 16 May 2021; Received in revised form 9 July 2021; Accepted 15 July 2021
Available online 22 July 2021
0968-0896/© 2021 Elsevier Ltd. All rights reserved.

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
hematologic malignancies such as chronic lymphocytic leukemia (CLL),
and follicular lymphoma (FL), was approved by the United States Food
and Drug Administration (FDA) in July 2014.35 However, the U.S. pre-
scribing information contains a black box warning for unexpected
adverse effects, such as fatal and severe diarrhea or colitis, hepatotox-
icity, pneumonitis, and intestinal perforation.36–37 Nevertheless, 1
showed high response rates and extraordinary efficacy in patients with
hematologic malignancies who were difficult to treat with conventional
anticancer drugs38 and demonstrated a tolerable safety profile in clinical
trials.
SUDHL-5 and MOLT-4, to determine the anti-cancer effects of the
compounds 6a-m and 8a-c (Fig. 3). Each compound showed a similar
pattern in the two cell lines, but higher efficacy was detected in SUDHL-
5, a DLBCL cell line that was used as a therapeutic target indication of 1,
while the efficacy was modest in MOLT-4, a T-ALL cell line. However,
only compound 6m showed the best anti-proliferative activity regard-
less of cell lines, which is consistent with results demonstrating the
strongest PI3Kδ inhibitory effect. Specifically, the CelTiter-Glo assay
showed a decrease in cell viability by approximately 14–20% after 6m
treatment of both cell lines, demonstrating that the anticancer effect of
6m was approximately five-fold higher than 1 for SUDHL-5 as well as
MOLT-4. In addition, we also found that several compounds, such as 6h
and 6j, as well as 8c, which showed excellent kinase activity, exhibit a
stronger anticancer effect than 1, although it was not completely
consistent with their kinase efficacy. Taken together, these results
demonstrated that most of the compounds showed enhanced efficacy in
vitro compared with 1. In particular, 6m was identified as the strongest
compound that inhibits the kinase activity of PI3Kδ, resulting in an
anticancer effect of more than 80% in vitro in both DLBCL and T-ALL.
The four isoforms of the catalytic subunit of PI3K share the same
To overcome these adverse effects of compound 1, we developed a
potent PI3Kδ inhibitor by specific structural modifications to lower the
toxicity associated with enhanced PI3Kδ inhibition and expanded the
target indications, such as T-ALL, which has yet to be treated with 1
(Fig. 2). These modifications including the substituent change of qui-
nazolinone moiety at C-5 or 6 positions (region A) and alteration of
chain (region B) into a hydrophobic pocket are mainly focused on
developing potent PI3Kδ inhibitors since the purinyl group as the hinge
binder and the phenyl group that affects the potency and selectivity
across the PI3K class I family are key pharmacophores.39
domain composition, but PI3K α/β subunits are ubiquitously expressed,
whereas PI3K δ/γ subunits are mainly restricted to the hematopoietic
cell system. In particular, PI3Kδ and PI3Kγ are enriched in immune cells
and regulate the development and function of innate and adaptive im-
munity. Recent reports8,40 raise concerns about the risk of potential
adverse effects such as autoimmune disease, a chronic inflammatory
disorder, allergies, and immunodeficiency in the case of poor hetero-
tropic specificity of inhibitors to pharmacological targets, due to the
interaction between p110δ and p110γ. Therefore, appropriate isomeric
selectivity (ratio of γ/δ) should be considered in developing small-
molecule inhibitors of PI3Kδ. In addition, preceding studies showed
2. Results and discussion
2.1. Pharmacological evaluation
2.1.1. Cell viability and enzyme-based assays in vitro
Fifteen effective compounds were synthesized by structural modifi-
cation of compound 1. Among them, an in vitro kinase assay based on
ATP site-dependent competitive binding of PI3Kδ was initially per-
formed to select potent PI3Kδ inhibitors. The concentrations for half-
maximal inhibition (IC50) of each compound were calculated. As
shown in Figure 3 and Table 1, the IC50 values of the compounds were in
the nanomolar (nM) range of 0.09–3.23. Among them, about half of the
compounds (6b, 6f, 6h, 6i, 6j, 6m, and 8a) exhibited excellent effi-
ciencies with sub-nanomolar functional potency when compared with
the reference PI3Kδ inhibitor, 1.
that 1 has substantially higher selectivity for PI3Kδ than for PI3Kα and
PI3Kβ, but not for PI3Kγ. Furthermore, Duvelisib, which demonstrated
simultaneously inhibitory ability toward PI3Kδ and PI3Kγ, was intro-
duced as a highly promising drug for the treatment of blood cancer and
autoimmune diseases, but its drug development scenario was similar to
that of 1 for almost similar reasons. The selectivity of this drug for PI3Kδ
Cell viability was analyzed in two blood cancer cell lines such as
Fig. 1. Representative structures of PI3K inhibitors. (A) pan-PI3K inhibitors. (B) Isoform-selective PI3K inhibitors. (C) Dual PI3K/mTOR inhibitors.
2

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
Fig. 2. Design of quinazolinone derivatives as PI3Kδ inhibitors based on 1.
Fig. 3. Anticancer activity of quinazolinone derivatives against two hematological malignant cell lines (SUDHL-5 and MOLT-4). The anti-proliferative activity of
SUDHL-5 and MOLT-4 was measured by CellTiter-Glo Luminescent Cell Viability Assay, and a p110δ enzymatic inhibitory activity assay was conducted using the
SelectScreenTM Biochemical Kinase Profiling Service. SUDHL-5 cell was treated with 10 µM of each compound and MOLT-4 cell was treated with 20 µM of each
compound. Each IC50 value is representative data.
and PI3Kγ is poor compared with the inhibitory efficacy for PI3Kβ.
Therefore, it is necessary to develop new drugs that can selectively
inhibit PI3Kδ, at least, rather than 1.
was evaluated, it was confirmed that they have excellent PI3K selec-
tivity, which means that the off-target effect on other kinases will be
very low. Therefore, based on the inhibition results, 6f and 6m were
ultimately selected and used for the following experiments including
various kinases inhibitory activity (Supporting Table 1).
Table 1 shows the SAR of quinazolinone derivatives intended for the
development of novel PI3Kδ-specific inhibitors with improved selec-
tivity for PI3Kδ over 1. As the size of the R1 and R2 groups increased, the
activity and selectivity tended to improve (Table 1 and Fig. 4). Thus, due
to the structure of their binding pocket, lipophilic R1 and cyclic group R2
were functionally better than hydrophilic and linear groups, respec-
tively. Similarly, the longer the side chain R2, the better the activity.
However, its activity decreased sharply if the length of the side chain of
R2 exceeded four carbons, such as a butyl group. Therefore, it was
appropriate for the side chain length of R2 to remain within the butyl
group. 6e was the best in p110 γ/δ selectivity but was excluded because
its affinity to p110δ was lower than 1 (IC50 = 1.82 nM, γ/δ ratio =
171.97). Although poor in isomeric selectivity, generally 6m showed
remarkable inhibitory potency with an IC50 value of 0.09 nM and
selectivity to p110δ, and 6f appeared to exhibit the second-highest po-
tency with an IC50 of 0.39 nM. Subsequently, when these two com-
pounds, 6m, and 6f, were selected and the activity for various kinases
2.1.2. Docking model
The strong affinity of 6f and 6m for PI3Kδ was once again explained
by the docking results, in which the purinyl group established hydrogen
bonding with Glu826 and Val828 at a distance of 2.9 Å as the hinge
binder, which was critical for the activity and stability of the PI3Kδ in-
hibitor (Fig. 4). In addition, docking analysis showed that the quinazo-
linone moiety forms a hydrophobic interaction with Ile777, Trp760, and
Met752, and a water-mediated hydrogen bonding bridge with Asp911.
The cyclopropyl and phenyl groups showed hydrophobic interaction
with Met752, Met900, and Thr833 and electrostatic interaction with
Asp832, Met900, and Thr833 in the hydrophobic pocket. These findings
suggest that the activity and selectivity of PI3Kδ are increased by posi-
tioning a substituent of an appropriate size in the hydrophobic pocket
due to competitive binding in the ATP-binding pocket for interaction
3

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
was remarkably inhibited by treatment with 6f and 6m. Next, we
evaluated the ability of 6f and 6m to induce apoptotic cell death.
As shown in Fig. 6B, the rate of cell death increased with the treat-
ment using 6f and 6m in both early and late stages of apoptosis.
Compared to 1, the compounds 6f and 6m were substantially more
powerful in inducing apoptosis of MOLT-4, while only 6m was excellent
in SUDHL-10. These results indicate that 6f and 6m are potent inhibitors
of PI3Kδ and remarkably suppress signal transduction during cancer cell
proliferation and survival. Subsequently, the antitumor activity of
compounds 6f and 6m was evaluated in vivo using a xenograft model. To
evaluate the antitumor effects of 6f and 6m in nude mice transplanted
with MOLT-4, 6f and 6m including 1 were administered once a day and
the variation in tumor mass was measured for 57 days. Oral adminis-
tration to mice of 10 mg/kg of 6f and 6m led to a tumor growth inhi-
bition rate (IR) of 55.1% for 6f and 56.7% for 6m, similar to reference
substance 1. It was also found that the three compounds do not induce
body weight loss. In conclusion, 6f and 6m are effective in suppressing
tumor growth in MOLT-4, although no significant improvement
compared to 1 was detected in the xenograft model (Fig. 6C).
Table 1
PI3K isoform inhibitory activity for purinyl-quinazolinone derivatives 6a-6m,
8a-8c
.
Comp.
No.
R1
R2
IC50 (nM)a
Ratio
(γ/δ)
p110
α
p110β
p110γ
p110δ
1 (6c)
6a
6b
6d
6e
5-F
6-F
5-F
5-F
5-F
5-F
5-F
5-Cl
5-Cl
5-Cl
5-Cl
5-CF3
5-
Et
409
295
152
385
878
222
879
301
356
205
490
326
134
613
27.4
29.2
15.3
72.2
313.0
19.1
155
1.08
1.09
0.50
1.08
1.82
0.39
2.05
0.50
0.50
0.60
1.70
3.23
0.09
25.37
26.78
30.60
66.85
171.97
48.97
75.60
36.60
125.00
17.33
37.52
17.43
95.55
cPr
Me
nPr
iPr
200
145
440
1670
199
6f
cPr
cBu
nPr
iPr
6g
6h
6i
854
129
18.3
62.5
10.4
63.8
56.3
8.6
1040
87.6
470
6j
cPr
cBu
nPr
cPr
6k
6l
>1000
162
2.1.4. In vivo PK study
6m
The pharmacokinetic profile was further evaluated to determine the
pharmacological potential of 6f and 6m (Fig. 7). Treatment with an oral
dose of 3 mg/kg of 6f yielded an area under the curve (AUClast) and Cmax
of 255.00 h*ng/mL and 41.89 ng/mL, respectively. The median Tmax
and t1/2 were 1 h and 7.7 h, respectively. These values were improved
overall compared to 1 under the same dose (AUClast = 201.88 h*ng/mL,
CH3
5-
8a
8b
8c
Me
Et
127
475
369
250
287
562
7.5
0.40
1.30
1.07
18.75
19.00
23.08
NH2
5-
24.7
24.7
NH2
5-
cPr
NH2
a
IC50 values of enzyme activity have been taken as a mean from three inde-
pendent experiments.
with amino acid residues outside the ATP-binding pocket (Fig. 5).
2.1.3. Cell-based in vitro study and tumor growth inhibition: A xenograft
model
PI3K catalyzes the conversion of PIP2 to PIP3, which induces phos-
phorylation at Ser473 and Thr308 of AKT, leading to signaling cascades
of downstream effectors. Activated AKT controls diverse cellular events
such as cell proliferation, protein synthesis, and survival via mTORC1,
which is upstream of 4EBP and S6. To investigate the effects of the
primary screened compounds 6f and 6m on the signal transduction of
PI3K, changes in the expression of key downstream factors were
analyzed by western blot in SUDHL-5 and MOLT-4 after treatment with
the compounds (Fig. 6A). Consistent with the inhibitory potency to
PI3Kδ, treatment with 6f and 6m demonstrated dramatic inhibition of
the expression of phospho-AKT at S473 and T308 compared to 1 in both
cell lines. Also, the activation of its downstream targets, S6 and 4EBP1,
Fig. 5. Space-filling representation and structure overlay of 1 (green) and 6f
(orange) bound to PI3Kδ active site.
Fig. 4. Predicted binding modes of compounds 6f (orange, Fig. 4A) and 6m (yellow, Fig. 4B) with PI3Kδ [PDB code: 4XE0]. Compound 1 is shown in green.
Hydrogen bonds are shown as green dashed lines. Key residues, as well as water molecules with the compounds, are indicated.
4

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
Fig. 6. Compounds 6f and 6m induce apoptosis-mediated cell death via inhibition of p-AKT activation and block tumor growth in the xenograft model. (A) SUDHL-5
and MOLT-4 cells were treated with 1 M and 5 M of each compound for 3 h, respectively. Western blot was performed to detect the phosphorylation levels of AKT
S473/T308, S6, and 4EBP1. β-actin was used as a loading control. (B) Flow cytometry was performed to measure apoptosis with annexin-V/propidium iodide (PI)
μ
μ
double staining. Cells were treated with 20 μM of each compound for 24 h, and then apoptotic cell death was measured. (C) In vivo antitumor efficacy in the MOLT-4
xenograft model. Tumor volume (TV) and changes in average body weight were measured after treatment with each compound at an oral dose of 10 mg/kg for
57 days.
Cmax = 53.02 ng/mL, Tmax = 0.5 h, t1/2 = 2.7 h). In contrast to 6f, and
contrary to our expectations, the pharmacokinetic parameters of 6m
from commercially available anthranilic acids 2a-e and amino acids 3a-
f, as shown in Scheme 1. Quinazolinone intermediate compounds 4a-m
were synthesized in one pot by reacting appropriate anthranilic acid 2
and the corresponding Boc-amino acid 3 with diphenyl phosphite and
pyridine, followed by the addition of aniline. Boc-deprotection of 4 with
trifluoroacetic acid (TFA) in dichloromethane yielded the primary
amines 5. Finally, the targeted derivatives 6a-m were synthesized by
reacting 6-chloropurine with amine 5 in the presence of trimethylamine
(TEA) and tert-butanol (t-BuOH).41,42
were very poor (AUClast = 37.24 h*ng/mL, Cmax = 11.06 ng/mL, Tmax
=
0.5 h, t1/2 = 8.3 h). Accordingly, the results indicate poor metabolic
stability of 6m compared with 6f.
2.1.5. Cytotoxicity
Investigation of toxicity against normal cells is an important aspect
to determine the safety profile of the drug molecule. Hence the potent
compounds 6f and 6m including standard compound 1(6c) were
examined for their toxicity. To determine the potential cytotoxicity of
these compounds (6f and 6m), we carried out an MTT assay with an
MRC-5 cell line. As shown in Table 2, none of the compounds showed a
high level of cytotoxicity. These results implied that 6f was an effective
and safe drug candidate for the treatment of hematologic malignancies.
Quinazolinone derivatives 8a-c (Table 1) containing amine group on
5th position was synthesized starting from the appropriate fluoro com-
pounds (Scheme 2). Nucleophilic attack of 4-methoxybenzylamine on
the appropriate fluoro compound (6b, 6c, and 6f) in the presence of TEA
in ethanol (EtOH) yielded intermediate compound 7. Deprotection of 7
with TFA in dichloromethane furnished the amino-quinazolinone 8.27
3. Conclusion
2.2. Chemistry
In summary, we have discovered a series of potent and selective
Quinazolinone derivatives 6a-m (Table 1) were synthesized starting
5

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
quinazolinone PI3Kδ inhibitors. SAR studies revealed that fluoro or
methyl substitution in the quinazolinone ring and hydrophobic cyclo-
propyl groups contributes to the potency and selectivity of PI3Kδ inhi-
bition. Further studies were conducted with 6f and 6m, and both
exhibited nanomolar enzymatic potencies against PI3Kδ. In addition, 6f
and 6m exhibited target modulation in cancer cells in the western
blotting analysis of SUDHL-5 and MOLT-4 cell lines and therefore
increased cell apoptosis. In particular, 6m displayed high selectivity for
the enzyme PI3Kδ and enhanced efficacy for cell viability; however, the
PK results were poor compared to 1. Compound 6f demonstrated anti-
tumor efficacies in xenograft models, in which PI3K pathways were
adequately inhibited as indicated by the reduction of p-AKT, p-S6, and p-
4EBP1 in tumor tissues. Hence, compound 6f was selected as a pre-
clinical candidate, and future communications will detail additional
evaluation of this compound.
Fig. 7. Plasma concentration and pharmacokinetic parameters of compounds
6f and 6m. Plasma concentration versus time profile and pharmacokinetic
parameters following treatment with 1 and 3 mg/kg 6f and 6m administered P.
O. to Sprague-Dawley rats. Pharmacokinetic parameters include maximum
concentration (Cmax), an area under the concentration versus time curve
(AUClast), time of maximum concentration (Tmax), and half-life (t1/2). *P < 0.05
vs. control.
4. Material and methods
4.1. Biological method
4.1.1. Cell viability assay
Cell viability was analyzed according to the manufacturer’s in-
structions using CellTiter-Glo Luminescent Cell Viability Assay (Prom-
ega, Madison MI, USA). Cells were seeded into 96-well plates at a density
of 5x104 cells in a 100 µL culture medium. CellTiter-Glo reagent was
added at a 1:1 vol ratio. After mixing the contents for 2 min on a shaker
to induce cell lysis, the plates were incubated for 10 min at room tem-
perature, and then the luminescence signal was read on a plate reader
(BioTek Instruments, Inc., Winooski, VT, USA). Luminescence readings
were background-subtracted and normalized to control wells.
Table 2
Cytotoxicity of 6f and 6m against MRC-5 cell line.
Comp. No.
% of Inhibition 20 µM
IC50 (µM)
1 (6c)
6f
18.41 ± 1.15
3.93 ± 2.13
23.12 ± 5.23
>100
>100
6m
49.11 ± 3.23
4.1.2. Biochemical kinase assay
The enzymatic inhibitor activity assay was performed by Select-
ScreenTM Biochemical Kinase Profiling Service (SSBK; Thermo Fisher
Scientific) according to the manufacturer’s instructions. The IC50 value
of the compound was determined by 10-point titration with 1 µM final
concentration of the compound in an ATP-competitive binding experi-
mental manner. For the kinase reaction, the kinase reaction buffer (30
mM HEPES, 4X ATP solution, 2x substrate/kinase mixture) was added to
the wells and then incubated for 60 min. A detection solution consisting
of europium labeled anti-ADP antibody, Alexa Fluor™ 647 labeled ADP
tracer and EDTA (to stop the kinase reaction) was added to the assay
well. IC50 values were determined by calculating the release rate of ADP
formation from the analytical well. The emission ratio was calculated by
dividing the intensity of the tracer (receptor) emission by the intensity of
the Eu (donor) emission at 615 nm, as shown in the equation below.
Em ission Ratio = AlexaFluor647 Em ission (665 nm )/Europium Em ission
(615 nm )
4.1.3. Western blot assay
Scheme 1. Reagents and conditions: (a) (i) diphenyl phosphite, pyridine,
50 ◦C; (ii) aniline, 55 ◦C; 1 N HCl, pH = 7 (2 steps); (b) CF3COOH, CH2Cl2, rt;
ammonia, pH = 8–12; (c) 6-chloropurine, TEA, t-BuOH, 80 ◦C.
Cells were collected and washed twice with cold PBS, and then
proteins were obtained with cell lysis buffer (CST, Danvers, MA, USA).
The proteins were separated by 10–12% SDS-polyacrylamide gel
Scheme 2. Reagents and conditions: (a) 4-methoxybenzylamine, TEA, EtOH, 180 ◦C; (b) CF3COOH, CH2Cl2, rt; ammonia, pH = 7.
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Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
electrophoresis and then transferred onto PVDF membranes (Millipore,
Burlington, MA, USA). The antibodies used for Western blot were as
follows: p-AKT S473 (CST; 9271), p-AKT T308 (CST; 9275), p-S6 (CST;
5364), p-4E-BP1 (CST; 9451), and β-actin (CST; 3700). The analysis was
performed with Peroxidase-AffiniPure Goat Anti-Rabbit (Jackson
ImmunoResearch; 111-035-003) or Horse Anti-Mouse (CST; 7076) IgG
antibodies, using the ECL western blotting detection system from GE
Healthcare AmershamTM (GE Healthcare, Chalfont St. Giles, UK).
inert atmosphere of nitrogen. Nuclear magnetic resonance spectra (1H
and 13C NMR) were recorded on a Bruker Unit 400 (400 MHz for 1H and
100 MHz for 13C) and Unity Inova (500 MHz for 1H and 125 MHz for
13C) instrument with CDCl3, CD3OD, and DMSO‑d6 as a solvent and
residual CHCl3 (δ 7.26 ppm), CH3OH (δ 3.31 ppm), and DMSO (δ 2.50
ppm) as an internal standard for 1H and CDCl3 (δ 77.0 ppm), CD3OD (δ
49.0 ppm), and DMSO‑d6 (δ 39.5 ppm) as an internal standard for 13C.
Resonance patterns are reported with the notations s (singlet),
d (doublet), t (triplet), q (quartet), and m (multiplet). In addition, the
notation brs is used to indicate a broad signal. Coupling constants (J) are
reported in hertz (Hz). IR spectra were recorded on a JASCO FT/IR-4600
and reported as cm-1. Optical rotations were measured with a JASCO
4.1.4. Cell apoptosis analysis by flow cytometry
Apoptosis analysis was performed according to the manufacturer’s
recommended procedures using the Annexin V-FITC/PI apoptosis
detection kit (BD Biosciences, Franklin Lakes, NJ, USA). Cells were
seeded into 6-well plates and incubated with the compounds for 24 h.
The cells were then washed with cold PBS and stained with Annexin V-
FITC and PI for 15 min at room temperature. Apoptotic cells were
analyzed using a flow cytometer (BD FACSCalibur, Franklin Lakes, NJ,
USA).
P1020 polarimetry or JASCO P2000 polarimetry and are reported as [α Dt
]
(concentration g/100 mL, solvent). The melting point was recorded on
an Electrothermal IA9300 or Mettler Toledo MP90 and reported as ◦C.
Thin-layer chromatography was carried out using plates coated with
Kieselgel 60F254 (Merck). For flash column chromatography, E. Merck
Kieselgel 60 (230–400 mesh) was used. Low-resolution mass spectra
(LRMS) were recorded on an SQ Detector 2 mass spectrometer.
XBridge®-C18 column (250 × 4.6 mm) was used as the stationary phase.
High-Performance Liquid Chromatography (HPLC) was recorded on a
waters 2695 separations module/2996 PDA detector with acetonitrile
and water as eluent and a detection wavelength of 254 nm.
4.1.5. Animal tumor experiments (Xenograft model and survival rate)
All animal experiments were performed by Biotoxtech (Biotoxtech
co. ltd, Cheongju, South Korea) according to relevant ethical regula-
tions. For xenograft studies, MOLT-4 cells were harvested and 2 × 106
cells in 0.1 mL of D-PBS were implanted subcutaneously on the back of
BALB/c-nude mice. After tumor growth, mice were treated daily with
Idelalisib 1, 6f, and 6m at 10 mg/kg by oral gavage for 57 consecutive
days. Tumor growth was monitored by a caliper (CD-15CX, Mitutoyo,
Japan), and tumor volume was calculated according to the formula of ½
× length × width. To evaluate survival results, BALB/c hematological
malignancies mice were administered intravenously with Daudi cells.
The test group was divided into 3 groups with 15 animals each, with
negative control (0 mg/kg), a test substance group (8 mg/kg), and a
comparative group. The compounds were administered orally in a dose
of 8 mg/kg for 56 days in the test substance groups. During the obser-
vation period, clinical signs were observed once daily. The body weight
was measured once a week.
4.2.1. General procedure a for the preparation of compounds 4a-m.
Diphenyl phosphite (1.65 eq. or 5 eq) was added to a stirred solution
of the corresponding anthranilic acid 2 (1 eq.) and appropriate Boc-
amino acid 3 (1.2 eq.) in pyridine (6 vol) at room temperature under
◦
N2 gas. The reaction mixture was heated to 40–45 C and stirred for
4–18 h (in case of using 1.65 eq diphenyl phosphite) or 2–5 h (in using 5
eq diphenyl phosphite). And then, aniline (1.4 eq.) was added to the
reaction mixture and the resulting reaction mixture was heated to 55 ◦C
for 5–16 h (in case of using 1.65 eq diphenyl phosphite) or 2–4 h (in case
of using 5 eq diphenyl phosphite). The completion of the reaction was
monitored by TLC on silica (Hexane-EtOAc = 4:1). The mixture was
cooled to room temperature, quenched with water, and then extracted
with EtOAc (2x times). The combined organic extracts were washed
with 1 N HCl solution (to remove pyridine and aniline) and brine. The
organic layer was dried with MgSO4, filtered, and evaporated under
reduced pressure. The crude residue was subjected to flash silica gel
(230–400 mesh) column chromatography (Hexane-EtOAc gradient) to
afford the title compounds.
4.1.6. PK study
The pharmacokinetics were performed by ChemOn Inc. (ChemOn
Inc., Suwon, South Korea) according to relevant ethical regulations.
Sprague-Dawley rats were used in PK studies of Idelalisib 1, 6f, and 6m,
and these compounds were administered once a day to three mice per
group at 1 and 3 mg/kg. The time points for blood samples collected
were 0.25, 0.5, 1, 2, 4, 8, and 24 h after oral administration. After
centrifugation, plasma samples were extracted and analyzed by LC-MS/
MS. The PK parameters were calculated with PhoenixTM WinNonlin®
software Ver. 6.2. (Pharsight Corporation, Mountain View, USA).
4.2.1.1. (S)-tert-Butyl (cyclopropyl(6-fluoro-4-oxo-3-phenyl-3,4-dihydroq
uinazolin-2-yl)methyl)carbamate (4a).. Synthesized from general pro-
cedure A using compound 2a and 3e under diphenyl phosphite (1.65
eq). 22% yield of the product as light ivory solid; mp 75.6–78 ◦C;
[
α 2D0
]
ꢀ 35.2886(c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.90 (dd, J
4.1.7. MTT assay
= 8.4 Hz, 2.8 Hz, 1H), 7.74–7.71 (m, 1H), 7.60–7.46 (m, 4H), 7.39 (d, J
= 7.6 Hz, 1H), 7.32 (d, J = 7.6 Hz, 1H), 5.54 (d, J = 7.6 Hz, 1H),
4.24–4.20 (m, 1H), 1.41 (s, 9H), 1.12–1.10 (m, 1H), 0.41–0.38 (m, 2H),
0.28–0.22 (m, 1H), 0.07–0.02 (m, 1H); 13C NMR (100 MHz, CDCl3) δ:
161.55 (d, JC-F = 3.1 Hz), 161.00 (d, JC-F = 247 Hz), 156.28, 155.00,
143.91 (d, JC-F = 1.9 Hz), 135.93, 130.22, 129.60, 129.53 (d, JC-F = 28.9
Hz), 129.33, 128.50, 123.06 (d, JC-F = 23.9 Hz), 122.28 (d, JC-F = 8.4
Hz), 111.96 (d, JC-F = 23.5 Hz), 79.68, 53.60, 28.31, 15.23, 3.49, 2.46;
IR (neat) 3343, 2977, 1681, 1604, 1590, 1482, 1364, 1336, 1273, 1228,
1159, 1067, 1021, 948, 886, 833, 748, 697 cmꢀ 1; LRMS (ESI) m/z
calculated for C23H25FN3O3 [M + H]+ 410.18, found 409.93.
Cytotoxicity was determined using an MTT assay. MRC-5 cells were
purchased from Korea Cell Line Bank. The cells were plated in 96-well
culture plates and seeded with 2 × 106 cells/mL (cells/well density).
Next, the cells were treated with 100 µL/well of the compound and 1%
FBS EMEM and incubated for 48 h at 37 ◦C in a CO2 incubator. Further,
at respective time points, 150 µL/well of culture media contained 10
times MTT solution (10:1 vol ratio) was added into each well, and cells
were incubated in a CO2 incubator for 3 h at 37 ◦C. The medium was
removed and Formazan crystals formed by the mitochondrial dehydro-
genase activity of vital cells were dissolved using 100 µL of DMSO fol-
lowed by transfer in new 96 well plate. The absorbance was read at 540
nm using 630 nm as a reference wavelength on an ELISA reader.
4.2.1.2. (S)-tert-Butyl (1-(5-fluoro-4-oxo-3-phenyl-3,4-dihydroquinazolin
-2-yl)ethyl)carbamate (4b). Synthesized from general procedure A using
compound 2b and 3a under diphenyl phosphite (1.65 eq). 20% yield of
the product as feather-like white solid; mp 158.6–163.9 ◦C;
4.2. Chemistry
Commercially available reagents were used without additional pu-
rification unless otherwise stated. All reactions were performed under an
[
α 2D1.8
]
ꢀ 39.9600(c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.72–7.66
7

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
(m, 1H), 7.62–7.50 (m, 4H), 7.39–7.37 (m, 1H), 7.29–7.28 (m, 1H),
7.14–7.09 (m, 1H), 5.60 (d, J = 7.6 Hz, 1H), 4.54–4.50 (m, 1H), 1.40 (s,
9H), 1.25 (d, J = 6.8 Hz, 3H). 13C NMR (100 MHz, CDCl3) δ: 161.49 (d,
JC-F = 265.1 Hz), 159.15 (d, JC-F = 4.3 Hz), 154.82, 149.19, 135.51,
134.92 (d, JC-F = 10.3 Hz), 130.39, 129.69, 129.60, 128.95, 128.90,
128.58, 123.07 (d, JC-F = 4.2 Hz), 113.55 (d, JC-F = 20.6 Hz), 110.79 (d,
JC-F = 5.7 Hz), 79.77, 47.85, 28.37, 20.68; IR (neat) 3315, 2977, 1705,
1681, 1610, 1590, 1524, 1473, 1364, 1294, 1238, 1170, 1071, 1018,
864, 820, 765, 698 cmꢀ 1; LRMS (ESI) m/z calculated for C21H22FN3O3
[M + H]+ 384.16, found 383.88.
yield of the product as a colorless liquid; [α D
]
20 ꢀ 64.9000(c 0.5, CH2Cl2);
1H NMR (400 MHz, CDCl3) δ: 7.67 (td, J = 8 Hz, 5.2 Hz, 1H), 7.59–7.47
(m, 4H) 7.34–7.29 (m, 2H), 7.12–7.07 (m, 1H), 5.46 (d, J = 8.8 Hz, 1H),
4.53–4.49 (m, 1H), 2.71–2.62 (m, 1H), 1.88–1.64 (m, 6H), 1.42 (s, 9H);
13C NMR (100 MHz, CDCl3) δ: 161.43 (d, JC-F = 265.2 Hz), 159.15 (d, JC-
F = 4.4 Hz), 157.21, 155.16, 149.09, 135.45, 134.80 (d, JC-F = 9.9 Hz),
130.23, 129.60, 129.34, 129.15, 128.61, 123.13 (d, JC-F = 3.6 Hz),
113.44 (d, JC-F = 20.6 Hz), 110.72 (d, JC-F = 5.6 Hz), 79.65, 54.03,
39.01, 28.29, 24.60, 23.41, 17.80; IR (neat) 3416, 3061, 2974, 2867,
1693, 1618, 1589, 1491, 1473, 1365, 1320, 1294, 1232, 1163, 1038,
869, 818, 736, 698 cmꢀ 1; LCMS (ESI) m/z calculated for C24H26FN3O3
[M + H]+ 424.20, found 424.20.
4.2.1.3. (S)-tert-Butyl (1-(5-fluoro-4-oxo-3-phenyl-3,4-dihydroquinazolin
-2-yl)butyl)carbamate (4d). Synthesized from general procedure A using
compound 2b and 3c under diphenyl phosphite (1.65 eq). 38% yield of
4.2.1.7. (S)-tert-Butyl (1-(5-chloro-4-oxo-3-phenyl-3,4-dihydroquinazolin
-2-yl)butyl)carbamate (4h). Synthesized from general procedure A using
compound 2c and 3c under diphenyl phosphite (1.65 eq). 45% yield of
the product as a white solid; mp 113.8–117 ◦C; [α D
]
20 ꢀ 47.6933 (c 0.5,
1
CH2Cl2); H NMR (400 MHz, CDCl3) δ: 7.71–7.66 (m, 1H), 7.62–7.48
the product as pale yellow solid; mp 187.6–188.8 ◦C; [α D20
ꢀ 45.7533(c
]
(m, 4H), 7.40 (d, J = 7.2 Hz, 1H), 7.29 (s, 1H), 7.13–7.08 (m, 1H), 5.42
(d, J = 8.8 Hz, 1H), 4.51–4.47 (m, 1H), 1.60–1.58 (m, 1H), 1.52–1.46
(m, 1H), 1.42 (s, 9H), 1.27–1.25 (m, 1H), 1.15–1.08 (m, 1H), 0.62 (t, J =
7.2 Hz, 3H); 13C NMR (100 MHz, CDCl3) δ: 161.45 (d, JC-F = 265.5 Hz),
159.15 (d, JC-F = 4.2 Hz), 158.69, 155.33, 149.18, 135.43, 134.82 (d, JC-
0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.61–7.51 (m, 5H), 7.46 (t, J
= 4.8 Hz, 1H), 7.40 (d, J = 7.2 Hz, 1H), 7.28 (d, J = 7.2 Hz, 1H), 5.41 (d,
J = 8.8 Hz, 1H), 4.50–4.45 (m, 1H), 1.61–1.56 (m, 1H), 1.52–1.45 (m,
1H), 1.29–1.25 (m, 1H), 1.14–1.09 (m, 1H), 0.63 (t, J = 7.2 Hz, 3H); 13
C
F = 10.6 Hz), 130.32, 129.59, 129.41, 129.01, 128.57, 123.04 (d, JC-F
=
NMR (100 MHz, CDCl3) δ: 160.28, 158.33, 155.32, 149.61, 135.67,
134.58, 133.84, 130.32, 129.59, 129.55, 129.42, 129.01, 128.55,
126.47, 118.18, 79.62, 51.23, 36.69, 28.32, 18.53, 13.11; IR (neat)
3383, 3053, 2982, 2877, 1693, 1613, 1588, 1552, 1493, 1455, 1392,
1266, 1174, 951, 813, 743, 703 cmꢀ 1; LCMS (ESI) m/z calculated for
C23H26ClN3O3 [M + H]+ 428.17, found 428.35.
2.8 Hz), 113.42 (d, JC-F = 20.9 Hz), 110.74 (d, JC-F = 5.3 Hz), 79.63,
51.19, 36.77, 28.32, 18.55, 13.11; IR (neat) 3394, 3052, 2977, 2963,
2876, 1768, 1694, 1608, 1589, 1473, 1294, 1244, 1163, 1037, 817, 736,
698 cmꢀ 1; LRMS (ESI) m/z calculated for C23H26FN3O3 [M + H]+
412.20, found 412.22.
4.2.1.4. (S)-tert-Butyl (1-(5-fluoro-4-oxo-3-phenyl-3,4-dihydroquinazolin
-2-yl)-2-methylpropyl)carbamate (4e). Synthesized from general pro-
cedure A using compound 2b and 3d under diphenyl phosphite (1.65
eq). 32% yield of the product as a white solid; mp 78–80.2 ◦C;
4.2.1.8. (S)-tert-Butyl (1-(5-chloro-4-oxo-3-phenyl-3,4-dihydroquinazolin
-2-yl)-2-methylpropyl)carbamate (4i). Synthesized from general proced-
ure A using compound 2c and 3d under diphenyl phosphite (1.65 eq).
37% yield of the product as light yellow solid; mp 55.7 ◦C;
[ ]
α 2D0 ꢀ 12.9000(c 0.5, CH2Cl2); 1H NMR (500 MHz, CDCl3) δ: 7.93 (d, J
[
α 2D0
]
ꢀ 55.2933 (c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.70 (m,
1H), 7.60–7.48 (m, 4H), 7.34–7.28 (m, 2H), 7.09 (t, J = 9.2 Hz, 1H),
5.43 (d, J = 9.6 Hz, 1H), 4.41–4.38 (m, 1H), 2.05–1.98 (m, 1H), 1.43 (s,
9H), 0.84 (d, J = 6.4 Hz, 3H), 0.71 (d, J = 6.4 Hz, 3H); 13C NMR (100
MHz, CDCl3) δ: 161.25 (d, JC-F = 265.2 Hz), 158.98 (d, JC-F = 4.2 Hz),
157.64, 155.15, 148.84, 135.37, 134.71 (d, JC-F = 10.3 Hz), 130.08,
129.39, 129.18, 128.42, 123.00 (d, JC-F = 3.6 Hz), 113.32 (d, JC-F = 20.5
Hz), 110.56 (d, JC-F = 5.7 Hz), 79.39, 56.05, 32.11, 28.16, 19.89, 16.28;
IR (neat) 3771, 3525, 2979, 2964, 2874, 1698, 1590, 1474, 1391, 1294,
1244, 1168, 1038, 818, 776, 735, 698 cmꢀ 1; LRMS (ESI) m/z calculated
for C23H26FN3O3 [M + H]+ 412.20, found 412.35.
= 8 Hz, 1H), 7.88 (d, J = 7.5 Hz, 1H), 7.60–7.50 (m, 3H), 7.36–7.23 (m,
3H), 5.36 (d, J = 9.5 Hz, 1H), 4.43–4.40 (m, 1H), 2.03–1.98 (m, 1H),
1.42 (s, 9H), 0.84 (d, J = 6.5 Hz, 3H), 0.72 (d, J = 7 Hz, 3H); 13C NMR
(125 MHz, CDCl3) δ: 160.32, 157.48, 155.29, 149.43, 135.71, 134.58,
133.84, 130.26, 129.67, 129.52, 129.31, 129.27, 128.54, 126.58,
118.20, 79.60, 56.19, 32.23, 28.30, 20.02, 16.38; IR (neat) 3429, 3312,
2965, 2926, 1690, 1587, 1490, 1455, 1365, 1276, 1161, 947, 814, 697
cmꢀ 1; LCMS (ESI) m/z calculated for C23H26ClN3O3 [M + H]+ 428.17,
found 428.28.
4.2.1.9. (S)-tert-Butyl ((5-chloro-4-oxo-3-phenyl-3,4-dihydroquinazolin-
2-yl)(cyclopropyl)methyl)carbamate (4j). Synthesized from general pro-
cedure A using compound 2c and 3e under diphenyl phosphite (5 eq).
4.2.1.5. (S)-tert-Butyl (cyclopropyl(5-fluoro-4-oxo-3-phenyl-3,4-dihydroq
uinazolin-2-yl)methyl)carbamate (4f). Synthesized from general pro-
cedure A using compound 2b and 3e under diphenyl phosphite (1.65
eq). 31% yield of the product as a white solid; mp 76.3–78.8 ◦C;
77% yield of the product as
a
white solid; mp 168.5 ◦C;
[
α 2D0
]
ꢀ 248.6667(c 0.5, CH2Cl2); 1H NMR (500 MHz, CDCl3) δ: 7.63–7.60
[
α D
]
16.8 ꢀ 36.7400 (c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ:
(m, 2H), 7.59–7.48 (m, 3H), 7.46 (dd, J = 6.5 Hz, 3 Hz, 1H), 7.38 (d, J =
7.5 Hz, 1H), 7.33 (d, J = 7 Hz, 1H), 5.53 (d, J = 7.5 Hz, 1H), 4.20–4.17
(m, 1H), 1.41 (s, 9H), 1.10–1.19 (m, 1H), 0.41–0.38 (m, 2H), 0.28–0.24
(m, 1H), 0.07–0.04 (m, 1H); 13C NMR (125 MHz, CDCl3) δ: 160.31,
157.61, 154.96, 149.63, 135.78, 134.59, 133.85, 130.15, 129.63,
129.57, 129.23, 128.57, 126.51, 118.19, 79.70, 53.66, 28.29, 15.05,
3.52, 2.41; IR (neat) 3728, 3704, 3622, 3595, 3385, 3055, 2975, 2931,
1693, 1610, 1587, 1491, 1455, 1365, 1165, 960, 813, 780, 737, 701
cmꢀ 1; LRMS (ESI) m/z calculated for C23H24ClN3O3 [M + H]+ 426.16,
found 426.22.
7.72–7.76 (m, 1H), 7.59–7.50 (m, 4H), 7.39–7.37 (m, 1H), 7.33–7.32
(m, 1H), 7.11 (t, J = 9.2 Hz, 1H), 5.54 (d, J = 8 Hz, 1H), 4.22–4.18 (m,
1H), 1.41 (s, 9H), 1.11–1.10 (m, 1H), 0.42–0.38 (m, 2H), 0.29–0.24 (m,
1H), 0.07–0.03 (m, 1H); 13C NMR (100 MHz, CDCl3) δ: 161.48 (d, JC-F
=
265.3 Hz), 159.17 (d, JC-F = 4.5 Hz), 157.97, 154.97, 149.23, 135.59,
134.82 (d, JC-F = 10 Hz), 130.15, 129.62, 129.60, 129.22, 128.61,
123.11 (d, JC-F = 3.4 Hz), 113.46 (d, JC-F = 20.9 Hz), 110.79 (d, JC-F
=
5.7 Hz), 79.70, 53.66, 28.30, 15.13, 3.52, 2.43; IR (neat) 3389, 2975,
1690, 1618, 1605, 1587, 1491, 1472, 1390, 1364, 1320, 1230, 1159,
1037, 882, 816, 763, 696 cmꢀ 1; LRMS (ESI) m/z calculated for
C23H24FN3O3 [M + H]+ 410.18, found 409.99.
4.2.1.10. (S)-tert-Butyl ((5-chloro-4-oxo-3-phenyl-3,4-dihydroquinazolin-
2-yl)(cyclobutyl)methyl)carbamate (4k). Synthesized from general pro-
cedure A using compound 2c and 3f under diphenyl phosphite (5 eq).
4.2.1.6. (S)-tert-Butyl (cyclobutyl(5-fluoro-4-oxo-3-phenyl-3,4-dihydroqui
nazolin-2-yl)methyl)carbamate (4g). Synthesized from general proced-
ure A using compound 2b and 3f under diphenyl phosphite (5 eq). 80%
95% yield of the product as a white solid; mp 61.5 ◦C; [α D
]
20 ꢀ 630.1333(c
0.5, CH2Cl2); 1H NMR (500 MHz, CDCl3) δ: 7.62–7.50 (m, 5H), 7.46 (t, J
8

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
= 4.5 Hz, 1H), 7.34–7.30 (m, 2H), 5.45 (d, J = 9 Hz, 1H), 4.51–4.48 (m,
1H), 2.69–2.64 (m, 1H), 1.87–1.64 (m, 6H), 1.42 (s, 9H); 13C NMR (125
MHz, CDCl3) δ: 160.30, 156.86, 155.15, 149.51, 135.67, 134.55,
133.83, 130.25, 129.62, 129.58, 129.36, 129.13, 128.58, 126.56,
118.16, 79.65, 54.04, 38.94, 28.29, 24.59, 23.38, 17.81; IR (neat) 3733,
3692, 3623, 3596, 3423, 3062, 2974, 1693, 1587, 1491, 1455, 1365,
1249, 1164, 1057, 951, 814, 776, 738, 696 cmꢀ 1; LRMS (ESI) m/z
calculated for C24H26ClN3O3 [M + H]+ 440.17, found 439.96.
cmꢀ 1; LRMS (ESI) m/z calculated for C18H16FN3O [M + H]+ 310.13,
found 309.97.
4.2.2.2. (S)-2-(1-Aminoethyl)-5-fluoro-3-phenylquinazolin-4(3H)-one
(5b). Synthesized from general procedure B using compound 4b. 91%
yield of the product as
a
white solid; mp 164.2–168.3 ◦C;
[
α 2D2.3
]
ꢀ 8.8000(c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.68 (td, J
= 8 Hz, 5.6 Hz, 1H), 7.57–7.50 (m, 4H), 7.28–7.26 (m, 2H), 7.09 (dd, J
= 10.4 Hz, 8 Hz, 1H), 3.68 (q, J = 6.4 Hz, 1H), 1.87 (brs, 2H), 1.27 (d, J
= 6.4 Hz, 3H); 13C NMR (100 MHz, CDCl3) δ: 161.38 (d, JC-F = 264.9
4.2.1.11. (S)-tert-Butyl (1-(4-oxo-3-phenyl-5-(trifluoromethyl)-3,4-dihy-
droquinazolin-2-yl)butyl)carbamate (4l). Synthesized from general pro-
cedure A using compound 2e and 3c under diphenyl phosphite (5 eq).
80% yield of the product as light yellow solid; mp 71.2 ◦C;
Hz), 161.96, 159.32 (d, JC-F = 4.4 Hz), 149.42, 136.01, 134.80 (d, JC-F
=
10.3 Hz), 129.98, 129.77, 129.46, 128.78, 128.25, 123.05 (d, JC-F = 4.2
Hz), 113.25 (d, JC-F = 20.5 Hz), 110.57 (d, JC-F = 5.6 Hz), 48.51, 23.29;
IR (neat) 3361, 3299, 2971, 1677, 1589, 1474, 1454, 1324, 1229, 1066,
904, 819, 758, 697 cmꢀ 1; LRMS (ESI) m/z calculated for C16H14FN3O
[M + H]+ 284.11, found 284.20.
[
α 2D0
]
ꢀ 35.9667(c 0.5, CH2Cl2); 1H NMR (500 MHz, CDCl3) δ: 7.92 (d, J
= 8 Hz, 1H), 7.87 (d, J = 7.5 Hz, 1H), 7.80 (t, J = 8 Hz, 1H), 7.62–7.50
(m, 3H), 7.42 (d, J = 7 Hz, 1H), 7.29 (d, J = 7.5 Hz, 1H), 5.40 (d, J = 9
Hz, 1H), 4.52–4.49 (m, 1H), 1.62–1.60 (m, 1H), 1.53–1.47 (m, 1H), 1.42
(s, 9H), 1.29–1.27 (m, 1H), 1.16–1.10 (m, 1H), 0.63 (t, J = 7.5 Hz, 3H);
13C NMR (125 MHz, CDCl3) δ: 159.12, 159.07, 155.36, 149.64, 135.56,
133.31, 132.27, 130.43, 129.65, 129.57, 129.29 (q, JC-F = 33 Hz),
128.89, 128.50, 126.17 (q, JC-F = 7.2 Hz), 123.17 (q, JC-F = 271.6 Hz),
118.36, 79.71, 51.23, 36.65, 28.32, 18.57, 13.08; IR (neat) 3735, 3595,
3054, 2964, 2932, 1698, 1590, 1490, 1367, 1307, 1156, 952, 830, 737,
698 cmꢀ 1; LRMS (ESI) m/z calculated for C24H26F3N3O3 [M + H]+
462.20, found 462.09.
4.2.2.3. (S)-2-(1-Aminobutyl)-5-fluoro-3-phenylquinazolin-4(3H)-one
(5d). Synthesized from general procedure B using compound 4d. 87%
yield of the product as colorless oil; [α D20
]
ꢀ 5.4867(c 0.5, CH2Cl2); 1H
NMR (400 MHz, CDCl3) δ: 7.67 (td, J = 8.4 Hz, 5.2 Hz, 1H), 7.57–7.48
(m, 4H), 7.28–7.25 (m, 2H), 7.08 (ddd, J = 10.4 Hz, 8.4 Hz, 0.8 Hz, 1H),
3.51 (dd, J = 8.4 Hz, 4.8 Hz, 1H), 1.73–1.65 (m, 3H), 1.50–1.42 (m, 1H),
1.38–1.28 (m, 1H), 1.18–1.09 (m, 1H), 0.69 (t, J = 7.2 Hz, 3H); 13C NMR
(100 MHz, CDCl3) δ: 161.51, 161.43 (d, JC-F = 265.1 Hz), 159.34 (d, JC-F
= 4.3 Hz), 149.49, 136.10, 134.73 (d, JC-F = 10.4 Hz), 129.95, 129.65,
129.38, 129.05, 128.23, 123.08 (d, JC-F = 4.2 Hz), 113.17 (d, JC-F = 20.7
Hz), 110.60 (d, JC-F = 5.8 Hz), 52.39, 39.15, 19.05, 13.39; IR (neat)
3372, 3051, 2962, 2873, 1689, 1586, 1472, 1394, 1295, 1274, 1180,
1036, 817, 749, 698 cmꢀ 1; LRMS (ESI) m/z calculated for C18H18FN3O
[M + H]+ 312.15, found 312.29.
4.2.1.12. (S)-tert-Butyl (cyclopropyl(5-methyl-4-oxo-3-phenyl-3,4-dihy-
droquinazolin-2-yl)methyl)carbamate (4m). Synthesized from general
procedure A using compound 2d and 3e under diphenyl phosphite (1.65
eq). 32% yield of the product as yellow solid; mp 155.3–165.8 ◦C;
[
α 2D0
]
ꢀ 33.0320(c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.62–7.48
(m, 5H), 7.38–7.32 (m, 2H), 7.23 (t, J = 6.8 Hz, 1H), 5.62 (d, J = 8 Hz,
1H), 4.23–4.19 (m, 1H), 2.82 (s, 3H), 1.41 (s, 9H), 1.12–1.10 (m, 1H),
0.40–0.37 (m, 2H), 0.30–0.25 (m, 1H), 0.08–0.04 (m, 1H); 13C NMR
(100 MHz, CDCl3) δ: 162.81, 156.33, 155.02, 148.76, 141.63, 136.29,
133.64, 130.20, 129.62, 129.57, 129.40, 129.25, 128.62, 125.43,
119.46, 79.53, 53.36, 28.29, 23.06, 15.15, 3.46, 2.32; IR (neat) 3376,
1980, 1669, 1590, 1499, 1472, 1364, 1226, 1161, 1045, 881, 810, 761,
691 cmꢀ 1; LRMS (ESI) m/z calculated for C24H27N3O3 [M + H]+ 406.21,
found 405.98.
4.2.2.4. (S)-2-(1-Amino-2-methylpropyl)-5-fluoro-3-phenylquinazolin-4
(3H)-one (5e). Synthesized from general procedure B using compound
4e. 77% yield of the product as a white solid; mp 159.6–160.4 ◦C; [α D20
+
]
33.0067(c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.64 (td, J = 8 Hz,
5.6 Hz, 1H), 7.56–7.47 (m, 4H), 7.28–7.26 (m, 2H), 7.06 (dd, J = 10 Hz,
8.4 Hz, 1H), 3.23 (d, J = 6.4 Hz, 1H), 2.05–1.97 (m, 1H), 1.63 (brs, 2H),
0.87 (d, J = 6.8 Hz, 3H), 0.73 (d, J = 6.8 Hz, 3H); 13C NMR (100 MHz,
CDCl3) δ: 161.17 (d, JC-F = 264.8 Hz), 160.66, 159.17 (d, JC-F = 4.3 Hz),
149.21, 136.11, 134.55 (d, JC-F = 10.5 Hz), 129.77, 129.37, 129.15,
129.10, 127.99, 122.96 (d, JC-F = 4.1 Hz), 112.94 (d, JC-F = 20.6 Hz),
110.36 (d, JC-F = 5.8 Hz), 58.03, 32.79, 20.23, 16.50; IR (neat) 3525,
3384, 3050, 2962, 2869, 1690, 1619, 1586, 1473, 1395, 1295, 1181,
1037, 818, 777, 733, 702 cmꢀ 1; LRMS (ESI) m/z calculated for
4.2.2. General procedure B for the preparation of compound 5a-m
To a stirred solution of the corresponding compound 4 (1 eq.) in
CH2Cl2 (0.085 M) was added trifluoroacetic acid (0.34 M) slowly at 0 ◦C
under N2 gas and stirred at room temperature for 1 h. The completion of
the reaction was monitored by TLC on silica (CH2Cl2-MeOH = 15:1) and
the reaction mixture was quenched with aq. ammonia (pH 8–12) solu-
tion at 0 ◦C. The reaction mixture was extracted with CH2Cl2. The
combined extracts were washed with water, dried over MgSO4, filtered,
and evaporated under reduced pressure. The residue was subjected to
flash column chromatography (CH2Cl2-MeOH gradient or EtOAc-MeOH
gradient) on silica gel to afford title compounds.
C
18H18FN3O [M + H]+ 312.15, found 312.29.
4.2.2.5. (S)-2-(Amino(cyclopropyl)methyl)-5-fluoro-3-phenylquinazolin-4
(3H)-one (5f). Synthesized from general procedure B using compound
4f. 99% yield of the product as pale yellow solid; mp 124.1–126.5 ◦C;
[
α 1D6.1
]
+ 9.24667(c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.68 (td,
J = 8 Hz, 5.6 Hz, 1H), 7.56–7.49 (m, 4H), 7.30–7.26 (m, 2H), 7.09 (dd, J
= 10 Hz, 8.4 Hz, 1H), 2.97 (d, J = 8.4 Hz, 1H), 1.79 (brs, 2H), 1.29–1.23
(m,1H), 0.53–0.38 (m, 2H), 0.14–0.02 (m, 2H); 13C NMR (100 MHz,
CDCl3) δ: 161.56 (d, JC-F = 230.4 Hz), 160.07, 159.29 (d, JC-F = 4.3 Hz),
149.53, 136.19, 134.73 (d, JC-F = 10.4 Hz), 129.87, 129.46, 129.39,
129.30, 128.16, 123.15 (d, JC-F = 4.2 Hz), 113.19 (d, JC-F = 20.5 Hz),
110.62 (d, JC-F = 5.7 Hz), 56.17, 17.92, 3.45, 3.07; IR (neat) 3357, 3056,
3006, 1714, 1678, 1621, 1583, 1473, 1319, 1226, 1038, 940, 897, 813,
762, 707 cmꢀ 1; LRMS (ESI) m/z calculated for C18H16FN3O [M + H]+
310.13, found 309.97.
4.2.2.1. (S)-2-(Amino(cyclopropyl)methyl)-6-fluoro-3-phenylquinazolin-4
(3H)-one (5a). Synthesized from general procedure B using compound
4a. 87% yield of the product as yellow syrup; [α D20
ꢀ 9.0157(c 0.5,
]
CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.89 (dd, J = 8.4 Hz, 2.8 Hz, 1H),
7.73 (dd, J = 9.2 Hz, 4.8 Hz, 1H), 7.57–7.46 (m, 4H), 7.31–7.26 (m, 2H),
2.99 (d, J = 8 Hz, 1H), 1.82 (brs, 2H), 1.30–1.25 (m, 1H), 0.52–0.40 (m,
2H), 0.12–0.01 (m, 2H); 13C NMR (100 MHz, CDCl3) δ: 161.58 (d, JC-F
=
3.3 Hz), 160.79 (d, JC-F = 246.6 Hz), 158.62, 144.12 (d, JC-F = 1.6 Hz),
136.47, 129.88, 129.67, 129.59, 129.50, 129.24 (d, JC-F = 34.9 Hz),
128.01, 122.92 (d, JC-F = 23.9 Hz), 122.03 (d, JC-F = 8.5 Hz), 111.71 (d,
JC-F = 23.5 Hz), 56.04, 17.95, 3.43, 3.06; IR (neat) 3370, 3066, 3005,
1676, 1588, 1482, 1344, 1271, 1226, 1161, 1019, 948, 833, 749, 697
4.2.2.6. (S)-2-(Amino(cyclobutyl)methyl)-5-fluoro-3-phenylquinazolin-4
(3H)-one (5g). Synthesized from general procedure B using compound
9

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
calculated for C19H18ClN3O [M + H]+ 340.12, found 340.19.
4g. 99% yield of the product as a colorless liquid; [α D20
+ 3.7666(c 0.5,
]
CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.66 (td, J = 8 Hz, 5.6 Hz, 1H),
7.60–7.47 (m, 4H), 7.30–7.26 (m, 2H), 7.08 (dd, J = 9.6 Hz, 8.8 Hz, 1H),
3.43 (d, J = 7.6 Hz, 1H), 2.81–2.71 (m, 1H), 1.99–1.94 (m, 1H),
4.2.2.11. (S)-2-(1-Aminobutyl)-3-phenyl-5-(trifluoromethyl)quinazolin-4
(3H)-one (5l). Synthesized from general procedure B using compound
4l. 99.6% yield of the product as a white solid; mp 136.2–136.8 ◦C;
1.86–1.58 (m, 7H); 13C NMR (100 MHz, CDCl3) δ: 161.38 (d, JC-F
=
α 2D0
]
ꢀ 156.1333(c 0.5, CH2Cl2); 1H NMR (500 MHz, CDCl3) δ: 7.92 (d, J
264.8 Hz), 159.96, 159.34 (d, JC-F = 4.3 Hz), 149.50, 136.26, 134.69 (d,
JC-F = 10.5 Hz), 129.98, 129.66, 129.38, 129.09, 128.26, 123.16 (d, JC-F
= 4.2 Hz), 113.17 (d, JC-F = 20.5 Hz), 110.61 (d, JC-F = 5.7 Hz), 56.80,
40.50, 24.90, 24.43, 17.71; IR (neat) 3370, 3061, 2968, 1857, 1688,
[
= 8 Hz, 1H), 7.85 (d, J = 7.5 Hz, 1H), 7.79 (t, J = 8 Hz, 1H), 7.56–7.48
(m, 3H), 7.27 (d, J = 7.5 Hz, 2H), 3.53 (dd, J = 8.5 Hz, 5 Hz, 1H), 1.81
(brs, 2H), 1.74–1.67 (m, 1H), 1.50–1.43 (m, 1H), 1.37–1.30 (m, 1H),
1.18–1.11 (m, 1H), 0.69 (t, J = 7.5 Hz, 3H); 13C NMR (125 MHz, CDCl3)
δ: 161.71, 159.31, 149.88, 136.14, 133.24, 132.33, 130.06, 129.81,
129.44, 129.13 (q, JC-F = 32.7 Hz), 128.89, 128.08, 125.98 (q, JC-F = 7.2
Hz), 123.18 (q, JC-F = 271.8 Hz), 118.12, 52.32, 39.07, 19.02, 13.36; IR
(neat) 3803, 3688, 3380, 3065, 2960, 2872, 1698, 1590, 1443, 1308,
1148, 956, 830, 698 cmꢀ 1; LRMS (ESI) m/z calculated for C19H18F3N3O
[M + H]+ 362.15, found 362.03.
1618, 1586, 1472, 1365, 1295, 1232, 1066, 1037, 818, 735, 698 cmꢀ 1
;
LRMS (ESI) m/z calculated for C19H18FN3O [M + H]+ 324.15, found
324.18.
4.2.2.7. (S)-2-(1-Aminobutyl)-5-chloro-3-phenylquinazolin-4(3H)-one
(5h). Synthesized from general procedure B using compound 4h. 54%
yield of the product as
a
white solid; mp 114.2–114.8 ◦C;
[
α 2D0
]
ꢀ 14.9933(c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 7.62–7.60
4.2.2.12. (S)-2-(Amino(cyclopropyl)methyl)-5-methyl-3-phenyl-
(m, 2H), 7.58–7.48 (m, 3H), 7.45 (dd, J = 6.4 Hz, 2.8 Hz, 1H), 7.28–7.25
(m, 2H), 3.50 (dd, J = 8 Hz, 4.4 Hz, 1H), 1.73–1.65 (m, 3H), 1.50–1.41
(m, 1H), 1.38–1.29 (m, 1H), 1.19–1.09 (m, 1H), 0.69 (t, J = 7.2 Hz, 3H);
13C NMR (100 MHz, CDCl3) δ: 161.15, 160.51, 149.91, 136.31, 134.47,
133.82, 129.97, 129.69, 129.42, 129.38, 129.05, 128.21, 126.54,
118.04, 52.43, 39.12, 19.05, 13.41; IR (neat) 3752, 3015, 2968, 2954,
2868, 1737, 1716, 1674, 1586, 1456, 1365, 1284, 1216, 1044, 810, 700
cmꢀ 1; LRMS (ESI) m/z calculated for C18H18ClN3O [M + H]+ 328.12,
found 328.20.
quinazolin-4(3H)-one (5m). Synthesized from general procedure B using
compound 4m. 77% yield of the product as pale yellow solid; mp
123.4–126.4 ◦C; [α D
]
20 ꢀ 4.2880(c 0.5, CH2Cl2); 1H NMR (400 MHz,
CDCl3) δ: 7.62–7.48 (m, 5H), 7.31–7.26 (m, 2H), 7.22–7.21 (m, 1H),
2.96 (d, J = 8.4 Hz, 1H), 2.82 (s, 3H), 1.89 (brs, 2H), 1.29–1.25 (m, 1H),
0.49–0.39 (m, 2H), 0.13–0.02 (m, 2H); 13C NMR (100 MHz, CDCl3) δ:
162.95, 158.82, 149.06, 141.47, 136.93, 133.58, 129.91, 129.47,
129.37, 129.17, 128.22, 125.47, 119.34, 56.08, 23.04, 17.97, 3.46,
3.10; IR (neat) 3371, 3009, 2968, 2923, 1715, 1677, 1565, 1472, 1375,
1285, 1134, 1041, 830, 765, 104 cmꢀ 1; LRMS (ESI) m/z calculated for
4.2.2.8. (S)-2-(1-Amino-2-methylpropyl)-5-chloro-3-phenylquinazolin-4
(3H)-one (5i). Synthesized from general procedure B using compound
4i. 36% yield of the product as a white solid; mp 94.5 ◦C in decompo-
C
19H19N3O [M + H]+ 306.15, found 305.96.
sition; [α 2D0
]
+ 15.5400(c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ:
4.2.3. General procedure C for the preparation of compound 6a-m
To a stirred solution of the corresponding amine 5 (1 eq.) in tert-
butanol (0.15 M) were added TEA (2 eq.) and 6-chloropurine (2 eq.) at
room temperature and the reaction mixture was refluxed for 10–27 h
under N2 gas. The completion of the reaction was monitored by TLC on
silica (CH2Cl2-MeOH = 20:1). After cooling to room temperature, the
reaction mixture was diluted with CH2Cl2 and washed with water until
6-chloropurine disappeared. The organic layer was dried over MgSO4,
filtered, and evaporated under reduced pressure. The residue was sub-
jected to flash column chromatography (CH2Cl2-MeOH gradient) on
silica gel to afford the title compound.
7.64–7.48 (m, 5H), 7.45 (dd, J = 6.4 Hz, 2.4 Hz, 1H), 7.29–7.25 (m, 2H),
3.24 (d, J = 6.4 Hz, 1H), 2.06–1.98 (m, 1H), 1.71 (brs, 2H), 0.87 (d, J =
6.8 Hz, 3H), 0.74 (d, J = 6.8 Hz, 3H); 13C NMR (100 MHz, CDCl3) δ:
160.54, 160.35, 149.77, 136.46, 134.43, 133.79, 129.97, 129.59,
129.41, 129.30, 129.28, 128.11, 126.59, 118.03, 58.22, 32.93, 20.40,
16.61; IR (neat) 3525, 3384, 3050, 2962, 2869, 1690, 1619, 1586, 1473,
1295, 1250, 1181, 1037, 818, 777, 702 cmꢀ 1; LRMS (ESI) m/z calcu-
lated for C18H18ClN3O [M + H]+ 428.12, found 328.20.
4.2.2.9. (S)-2-(Amino(cyclopropyl)methyl)-5-chloro-3-phenylquinazolin-
4(3H)-one (5j). Synthesized from general procedure B using compound
4j. 98% yield of the product as a white solid; mp 147.2–148.7 ◦C;
4.2.3.1. (S)-2-(((7H-Purin-6-yl)amino)(cyclopropyl)methyl)-6-fluoro-3-
phenylquinazolin-4(3H)-one (6a). Synthesized from general procedure C
for 24 h using compound 5a. 62% yield of the product as light ivory
[
α 2D0
]
ꢀ 43.3333(c 0.5, CH2Cl2); 1H NMR (500 MHz, CDCl3) δ: 7.65–7.60
(m, 2H), 7.56–7.46 (m, 4H), 7.31–7.26 (m, 2H), 2.97 (d, J = 8.5 Hz, 1H),
1.76 (brs, 2H), 1.29–1.23 (m, 1H), 0.52–0.40 (m, 2H), 0.13–0.03 (m,
2H); 13C NMR (125 MHz, CDCl3) δ: 160.51, 160.09, 149.98, 136.44,
134.51, 133.85, 129.94, 129.54, 129.50, 129.43, 129.35, 128.18,
126.64, 118.11, 56.26, 17.95, 3.52, 3.12; IR (neat) 3727, 3706, 3623,
3598, 3366, 3066, 2943, 1685, 1585, 1550, 1490, 1455, 1285, 1240,
957, 813, 777, 735, 699 cmꢀ 1; LRMS (ESI) m/z calculated for
solid; mp 168.3 ◦C; [α D16.4
]
+ 111.833(c 0.5, CH2Cl2); 1H NMR (400 MHz,
CDCl3) δ: 13.67 (brs, 1H), 8.30 (s, 1H), 7.98 (s, 1H), 7.89 (dd, J = 8 Hz,
2.4 Hz, 1H), 7.74–7.71 (m, 1H), 7.64–7.40 (m, 6H), 6.84–6.82 (m, 1H),
4.96 (m, 1H), 1.41–1.37 (m, 1H), 0.51–0.49 (m, 2H), 0.40–0.38 (m, 1H),
0.24–0.21 (m, 1H); 13C NMR (100 MHz, CDCl3) δ: 161.73 (d, JC-F = 3.3
Hz), 160.99 (d, JC-F = 247 Hz), 156.05, 153.60, 151.90, 149.78, 143.90
C
18H16ClN3O [M + H]+ 326.11, found 326.07.
(d, JC-F = 1.7 Hz), 138,45, 136.07, 129.85, 129.64, 129.60 (d, JC-F
35.4 Hz), 129.41, 128.81, 123.08 (d, JC-F = 23.8 Hz), 122.21 (d, JC-F
=
=
8.7 Hz), 119.35, 111.87 (d, JC-F = 23.5 Hz), 53.55, 15.32, 3.63, 3.10; IR
(neat) 3724, 3623, 3008, 1679, 1588, 1482, 1357, 1226, 946, 834, 747
cmꢀ 1; LRMS (ESI) m/z calculated for C23H18FN7O [M + H]+ 428.16,
found 427.97; HPLC purity 100% ee.
4.2.2.10. (S)-2-(Amino(cyclobutyl)methyl)-5-chloro-3-phenylquinazolin-
4(3H)-one (5k.. Synthesized from general procedure B using compound
4k. 99% yield of the product as a white solid; mp 80–82 ◦C;
[
α 2D0
]
ꢀ 39.6667(c 0.5, CH2Cl2); 1H NMR (500 MHz, CDCl3) δ: 7.59–7.50
(m, 5H), 7.43 (dd, J = 6 Hz, 2.5 Hz, 1H), 7.29 (d, J = 7.5 Hz, 2H), 3.41
(d, J = 8 Hz, 1H), 2.78–2.73 (m, 1H), 1.95–1.94 (m, 1H), 1.81–1.59 (m,
7H); 13C NMR (125 MHz, CDCl3) δ: 160.43, 159.61, 149.87, 136.44,
134.31, 133.71, 129.94, 129.64, 129.33, 129.29, 129.01, 128.18,
126.56, 117.97, 56.81, 40.40, 24.86, 24.36, 17.67; IR (neat) 3726,
3705, 3623, 3597, 3376, 3059, 2973, 2939, 1687, 1585, 1550, 1489,
1455, 1330, 1268, 1211, 951, 814, 774, 694, 671 cmꢀ 1; LRMS (ESI) m/z
4.2.3.2. (S)-2-(1-((7H-Purin-6-yl)amino)ethyl)-5-fluoro-3-phenyl-
quinazolin-4(3H)-one (6b). Synthesized from general procedure C for
36 h using compound 5b. 51% yield of the product as a white solid; mp
282.4–284 ◦C; [α D20
]
+ 189.8638(c 0.5, CH2Cl2). 1H NMR (400 MHz,
DMSO‑d6) δ: 12.93 (brs, 1H), 8.17 (m, 2H), 7.93 (d, J = 4.8 Hz, 1H), 7.79
(td, J = 8 Hz, 5.6 Hz, 1H), 7.67–7.66 (m, 1H), 7.58 (m, 2H), 7.49–7.47
10

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
(m, 2H), 7.30–7.26 (m, 1H), 4.82 (m, 1H), 1.46 (d, J = 6.4 Hz, 3H); 13
C
1H), 5.43 (m, 1H), 2.98–2.93 (m, 1H), 2.04–2.01 (m, 1H), 1.91–1.68 (m,
5H); 13C NMR (100 MHz, CDCl3) δ: 161.33 (d, JC-F = 265 Hz), 159.35 (d,
JC-F = 4.2 Hz), 157.01, 154.09, 151.87, 149.81, 149.09, 138.50, 135.60,
134.85 (d, JC-F = 10.3 Hz), 129.67, 129.45, 129.18, 129.02, 123.33 (d,
JC-F = 3.9 Hz), 119.39, 113.50 (d, JC-F = 20.5 Hz), 110.67 (d, JC-F = 5.7
Hz), 53.64, 39.25, 24.94, 24.04, 17.94; IR (neat) 3217, 3060, 2970,
2861, 1690, 1608, 1588, 1473, 1321, 1295, 1242, 1037, 1024, 933, 893,
818, 735, 698 cmꢀ 1; LRMS (ESI) m/z calculated for C24H20FN7O [M +
H]+ 442.18, found 442.04; HPLC purity 96.92% ee.
NMR (100 MHz, DMSO‑d6) δ: 160.54 (d, JC-F = 262 Hz), 159.83, 158.44
(d, JC-F = 3.9 Hz), 151.91, 149.14, 136.10, 135.29 (d, JC-F = 10.6 Hz),
129.33, 129.23, 129.10, 129.06, 128.88, 123.06 (d, JC-F = 2.7 Hz),
113.09 (d, JC-F = 20.5 Hz), 111.31 (d, JC-F = 5.4 Hz), 47.87, 18.89; IR
(neat) 3370, 3136, 2989, 2934, 1687, 1606, 1513, 1476, 1230, 1080,
1021, 830, 813, 753, 699 cmꢀ 1; LRMS (ESI) m/z calculated for
C
21H16FN7O [M + H]+ 402.14, found 402.18; HPLC purity 95.67% ee.
4.2.3.3. (S)-2-(1-((7H-Purin-6-yl)amino)butyl)-5-fluoro-3-phenyl-
quinazolin-4(3H)-one (6d). Synthesized from general procedure C for
15 h using compound 5d. 68% yield of the product as pale yellow solid;
4.2.3.7. (S)-2-(1-((7H-Purin-6-yl)amino)butyl)-5-chloro-3-phenyl-
quinazolin-4(3H)-one (6h). Synthesized from general procedure C for
16 h using compound 5h. 63% yield of the product as pale yellow solid;
mp 160.3 ◦C in decomposition; [α 2D0
]
+ 156.7200(c 0.5, CH2Cl2); 1H
mp 171.8–173.0 ◦C; [α D20
]
+ 187.3333(c 0.5, CH2Cl2); 1H NMR (400
NMR (500 MHz, CDCl3) δ: 8.34 (s, 1H), 7.97 (s, 1H), 7.65–7.55 (m, 5H),
7.45 (d, J = 8.5 Hz, 1H), 7.34 (d, J = 8 Hz, 1H), 7.06 (t, J = 9.5 Hz, 1H),
6.82 (d, J = 7.5 Hz, 1H), 5.27 (m, 1H), 1.81–1.79 (m, 2H), 1.40–1.32 (m,
1H), 1.22–1.18 (m, 1H), 0.65 (t, J = 7.5 Hz, 3H); 13C NMR (125 MHz,
CDCl3) δ: 161.37 (d, JC-F = 265.2 Hz), 159.39 (d, JC-F = 4.2 Hz), 158.69,
154.15, 152.02, 149.75, 149.21, 138.51, 135.63, 134.84 (d, JC-F = 10.2
Hz), 129.88, 129.61, 129.51, 128.96, 123.23 (d, JC-F = 4 Hz), 119.51,
113.42 (d, JC-F = 20.5 Hz), 110.71 (d, JC-F = 5.7 Hz), 50.80, 36.41,
18.74, 13.16; IR (neat) 3414, 3050, 2978, 2963, 2879, 1768, 1696,
MHz, CDCl3) δ: 13.82 (brs, 1H), 8.35 (s, 1H), 7.98 (s, 1H), 7.61–7.53 (m,
6H), 7.42 (dd, J = 6.4 Hz, 2.8 Hz, 1H), 7.36 (d, J = 8.4 Hz, 1H), 6.83 (d,
J = 8.4 Hz, 1H), 5.25 (m, 1H), 1.83–1.78 (m, 2H), 1.40–1.33 (m, 1H),
1.27–1.20 (m, 1H), 0.66 (t, J = 7.2 Hz, 3H); 13C NMR (100 MHz, CDCl3)
δ: 160.49, 158.31, 154.19, 152.02, 149.65, 138.47, 135.86, 134.47,
133.85, 129.87, 129.61, 129.57, 129.52, 128.95, 126.64, 118.16, 50.83,
36.35, 18.73, 13.16; IR (neat) 3545, 3399, 3255, 3051, 2962, 2873,
2824, 1686, 1611, 1585, 1551, 1265, 1175, 947, 814, 735, 699 cmꢀ 1
;
1611, 1590, 1474, 1396, 1324, 1266, 1244, 1180, 819, 737, 700 cmꢀ 1
;
LRMS (ESI) m/z calculated for C23H20ClN7O [M + H]+ 446.15, found
LRMS (ESI) m/z calculated for C23H20FN7O [M + H]+ 430.18, found
446.24; HPLC purity 100% ee.
430.02; HPLC purity 100% ee.
4.2.3.8. (S)-2-(1-((7H-Purin-6-yl)amino)-2-methylpropyl)-5-chloro-3-
phenylquinazolin-4(3H)-one (6i). Synthesized from general procedure C
for 36 h using compound 5i. 73% yield of the product as an ivory solid;
4.2.3.4. (S)-2-(1-((7H-Purin-6-yl)amino)-2-methylpropyl)-5-fluoro-3-
phenylquinazolin-4(3H)-one (6e). Synthesized from general procedure C
for 24 h using compound 5e. 46% yield of the product as a white solid;
mp 159.3 ◦C; [α 2D0
]
+ 139.5933(c 0.5, CH2Cl2); 1H NMR (400 MHz,
mp 217.2 ◦C; [α 2D0
]
+ 94.0267(c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3)
CDCl3) δ: 13.87 (brs, 1H), 8.31 (s, 1H), 7.99 (s, 1H), 7.62–7.53 (m, 5H),
7.45 (dd, J = 7.6 Hz, 0.8 Hz, 1H), 7.38–7.36 (m, 1H), 7.33–7.32 (m, 1H),
6.62 (d, J = 9.2 Hz, 1H), 5.29 (m, 1H), 2.32–3.24 (m, 1H), 0.97 (d, J =
6.8 Hz, 3H), 0.85 (d, J = 6.8 Hz, 3H); 13C NMR (100 MHz, CDCl3) δ:
160.48, 157.39, 154.36, 151.91, 149.50, 138.48, 135.89, 134.49,
133.91, 129.85, 129.73, 129.49, 129.42, 129.35, 128.98, 126.78,
119.49, 118.16, 55.46, 32.79, 20.06, 16.99; IR (neat) 3650, 3409, 3061,
2964, 2928, 1714, 1689, 1610, 1586, 1455, 1362, 1298, 1221, 951, 814,
737, 700 cmꢀ 1; LRMS (ESI) m/z calculated for C23H20ClN7O [M + H]+
446.15, found 446.24; HPLC purity 98.49% ee.
δ: 13.65 (brs, 1H), 8.30 (s, 1H), 7.98 (s, 1H), 7.67 (td, J = 8.4 Hz, 5.6 Hz,
1H), 7.62–7.51 (m, 4H), 7.38–7.33 (m, 2H), 7.12–7.07 (m, 1H), 6.64 (d,
J = 8.8 Hz, 1H), 5.31–5.29 (m, 1H), 2.32–2.23 (m, 1H), 0.98 (d, J = 6.8
Hz, 3H), 0.85 (d, J = 6.8 Hz, 3H); 13C NMR (100 MHz, CDCl3) δ: 161.39
(d, JC-F = 265 Hz), 159.38 (d, JC-F = 4.3 Hz), 157.73, 154.32, 151.93,
149.06, 138.48, 135.65, 134.91 (d, JC-F = 10.1 Hz), 129.87, 129.55,
129.42, 129.36, 128.99, 123.37 (d, JC-F = 4.3 Hz), 113.56 (d, JC-F = 20.6
Hz), 110.71 (d, JC-F = 5.5 Hz), 55.42, 32.88, 20.06, 17.03; IR (neat)
3734, 3649, 3418, 3065, 2965, 1691, 1608, 1473, 1294, 1232, 1038,
932, 818, 735, 698 cmꢀ 1; LRMS (ESI) m/z calculated for C23H20FN7O
[M + H]+ 430.18, found 430.32; HPLC purity 100% ee.
4.2.3.9. (S)-2-(((7H-Purin-6-yl)amino)(cyclopropyl)methyl)-5-chloro-3-
phenylquinazolin-4(3H)-one (6j). Synthesized from general procedure C
for 19 h using compound 5j. 72% yield of the product as pale yellow
4.2.3.5. (S)-2-(((7H-Purin-6-yl)amino)(cyclopropyl)methyl)-5-fluoro-3-
phenylquinazolin-4(3H)-one (6f). Synthesized from general procedure C
for 10 h using compound 5f. 46% yield of the product as a white solid;
solid; mp 169.7 ◦C; [α D20
]
+ 800.1333(c 0.5, CH2Cl2); 1H NMR (500 MHz,
CDCl3) δ: 13.90 (brs, 1H), 8.30 (s, 1H), 7.97 (s, 1H), 7.58–7.51 (m, 6H),
7.43–7.39 (m, 2H), 6.93 (d, J = 7 Hz, 1H), 4.87 (m, 1H), 1.38–1.37 (m,
1H), 0.51–0.49 (m, 2H), 0.38 (m, 1H), 0.23–0.22 (m, 1H); 13C NMR
(125 MHz, CDCl3) δ: 160.52, 157.43, 153.54, 151.86, 149.62, 138.52,
135.96, 134.44, 133.87, 129.67, 129.57, 129.33, 128.92, 126.69,
119.38, 118.12, 53.76, 15.08, 3.70, 3.11; IR (neat) 3728, 3706, 3623,
3600, 2919, 2850, 1691, 1614, 1588, 1456, 1286, 1269, 1247, 962, 777,
756, 735, 671 cmꢀ 1; LRMS (ESI) m/z calculated for C23H18ClN7O [M +
H]+ 444.13, found 444.12; HPLC purity 98.61% ee.
mp 183 ◦C; [α 1D8.3
]
+ 153.953(c 0.5, CH3OH); 1H NMR (400 MHz,
CD3OD) δ: 8.16 (s, 1H), 8.13 (s, 1H), 7.79–7.73 (m, 1H), 7.61–7.47 (m,
6H), 7.20–7.15 (m, 1H), 4.79–4.78 (m, 1H), 3.33–3.32 (m, 1H),
1.49–1.46 (m, 1H), 0.59–0.56 (m, 2H), 0.42 (m, 1H), 0.26–0.22 (m, 1H);
13C NMR (100 MHz, CD3OD) δ: 162.55 (d, JC-F = 263.2 Hz), 161.16 (d,
JC-F = 4.3 Hz), 159.44, 153.50, 150.77, 141.06, 137.33, 136.56 (d, JC-F
= 10.5 Hz), 130.89, 130.61, 130.48, 130.45, 124.41 (d, JC-F = 4 Hz),
114.50 (d, JC-F = 20.9 Hz), 111.54 (d, JC-F = 5.9 Hz), 55.74, 15.62, 4.18,
3.77; IR (neat) 3412, 3005, 2806, 1714, 1610, 1589, 1473, 1362, 1228,
1037, 818, 697 cmꢀ 1; LRMS (ESI) m/z calculated for C23H18FN7O [M +
H]+ 428.16, found 427.95; HPLC purity 100% ee.
4.2.3.10. (S)-2-(((7H-Purin-6-yl)amino)(cyclobutyl)methyl)-5-chloro-3-
phenylquinazolin-4(3H)-one (6k). Synthesized from general procedure C
for 36 h using compound 5k. 50% yield of the product as pale yellow
solid; mp 155.0–157.0 ◦C; [α D20
+ 640.0000(c 0.5, CH2Cl2); 13.37 (brs,
]
4.2.3.6. (S)-2-(((7H-Purin-6-yl)amino)(cyclobutyl)methyl)-5-fluoro-3-
phenylquinazolin-4(3H)-one (6g). Synthesized from general procedure C
for 33 h using compound 5 g. 35% yield of the product as light ivory
1H), 8.29 (s, 1H), 8.00 (s, 1H), 7.65–7.58 (m, 3H), 7.55–7.44 (m, 3H),
7.39–7.34 (m, 2H), 6.68 (d, J = 9.2 Hz, 1H), 5.40 (m, 1H), 2.97–2.91 (m,
1H), 2.05–2.01 (m, 1H), 1.90–1.67 (m, 5H); 13C NMR (100 MHz, CDCl3)
δ: 160.45, 156.66, 154.08, 151.82, 149.77, 149.53, 138.49, 135.84,
134.44, 133.87, 129.70, 129.68, 129.46, 129.43, 129.19, 129.01,
126.75, 119.27, 118.13, 53.73, 39.17, 24.94, 24.05, 17.95; IR (neat)
solid; mp 154.8 ◦C; [α D20
]
+ 73.1266(c 0.5, CH2Cl2); 1H NMR (400 MHz,
CDCl3) δ: 13.92 (brs, 1H), 8.28 (s, 1H), 7.98 (s, 1H), 7.69–7.63 (m, 1H),
7.59 (t, J = 7.6 Hz, 1H), 7.52–7.50 (m, 2H), 7.47–7.4 (m, 1H), 7.38 (d, J
= 8 Hz, 1H), 7.35 (m, 1H), 7.08 (t, J = 9.2 Hz, 1H), 6.79 (d, J = 8.8 Hz,
11

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
3725, 3703, 3623, 3600, 3414, 3057, 2969, 1714, 1612, 1587, 1455,
1363, 1222, 1092, 953, 815, 736, 701 cmꢀ 1; LRMS (ESI) m/z calculated
for C24H20ClN7O [M + H]+ 458.15, found 458.33; HPLC purity 96.17%
ee.
procedure D using compound 6c. 29% yield of the product as pale yel-
low solid; mp 145.8 ◦C; [α D20
]
+ 64.8079(c 0.5, CH2Cl2); 1H NMR (400
MHz, CDCl3) δ: 8.81–8.79 (m, 1H), 8.31 (s, 1H), 7.96 (s, 1H), 7.60–7.54
(m, 3H), 7.46–7.42 (m, 2H), 7.36–7.34 (m, 1H), 7.25–7.23 (m, 2H), 6.86
(d, J = 8 Hz, 1H), 6.81 (d, J = 8.8 Hz, 2H), 6.47 (d, J = 8.4 Hz, 1H), 5.13
(m, 1H), 4.32–4.30 (m, 2H), 3.76 (s, 3H), 1.98–1.93 (m, 1H), 1.81–1.78
(m, 1H), 0.86 (t, J = 7.2 Hz, 3H); 13C NMR (100 MHz, CDCl3) δ: 164.72,
158.78, 152.06, 150.46, 148.92, 136.05, 135.79, 130.24, 129.99,
129.57, 129.01, 128.63, 114.02, 113.07, 106.81, 105.07, 55.27, 53.43,
46.75, 27.62, 10.00; IR (neat) 3321, 3040, 2932, 2884, 2830, 1703,
1655, 1564, 1510, 1445, 1327, 1247, 1132, 1031, 814, 698 cmꢀ 1; LRMS
(ESI) m/z calculated for C30H28N8O2 [M + H]+ 533.23, found 533.37.
4.2.3.11. (S)-2-(1-((7H-Purin-6-yl)amino)butyl)-3-phenyl-5-(tri-
fluoromethyl)quinazolin-4(3H)-one (6l). Synthesized from general pro-
cedure C for 27 h using compound 5l. 76% yield of the product as a
white solid; mp 159.9 ◦C; [α D20
]
+ 193.5867(c 0.5, CH2Cl2); 1H NMR
(500 MHz, CDCl3) δ: 8.35 (s, 1H), 7.99 (s, 1H), 7.86 (d, J = 8 Hz, 1H),
7.83 (d, J = 7.5 Hz, 1H), 7.74 (t, J = 8 Hz, 1H), 7.60–7.53 (m, 4H), 7.36
(d, J = 8 Hz, 1H), 6.87 (d, J = 7 Hz, 1H), 5.28–5.27 (m, 1H), 1.82–1.81
(m, 2H), 1.42–1.35 (m, 1H), 1.24–1.19 (m, 1H), 0.65 (t, J = 7.5 Hz, 3H);
13C NMR (125 MHz, CDCl3) δ: 159.32, 159.02, 154.17, 151.99, 149.68,
138.55, 135.74, 133.29, 132.43, 129.95, 129.65, 129.14 (q, JC-F = 32.6
Hz), 128.87, 128.81, 126.18 (q, JC-F = 7.1 Hz), 123.17 (q, JC-F = 271.6
Hz), 119.51, 118.30, 50.84, 36.29, 18.76, 13.11; IR (neat) 3735, 3596,
3421, 3051, 2962, 2933, 2872, 1697, 1613, 1590, 1474, 1456, 1307,
1153, 951, 830, 737, 698 cmꢀ 1; LRMS (ESI) m/z calculated for
4.2.4.3. (S)-2-(((7H-purin-6-yl)amino)(cyclopropyl)methyl)-5-((4-
methoxybenzyl)amino)-3-phenylquinazolin-4(3H)-one (7c). Synthesized
from general procedure D using compound 6f. 31% yield of the product
as yellow solid; mp 153.8 ◦C; [α D16.3
]
+ 256.727(c 0.5, CH2Cl2); 1H NMR
(400 MHz, CDCl3) δ: 14.03 (brs, 1H), 8.83–8.81 (m, 1H), 8.31 (s, 1H),
7.97 (s, 1H), 7.58–7.39 (m, 6H), 7.24–7.22 (m, 2H), 7.03 (d, J = 8.4 Hz,
1H), 6.86 (d, J = 7.6 Hz, 1H), 6.80 (d, J = 7.6 Hz, 2H), 6.46 (d, J = 8 Hz,
1H), 4.90 (m, 1H), 4.31–4.30 (m, 2H), 3.74 (s, 3H), 1.39–1.38 (m, 1H),
0.48–0.39 (m, 3H), 0.23 (m, 1H); 13C NMR (100 MHz, CDCl3) δ: 164.69,
158.70, 155.56, 151.87, 150.39, 148.93, 138.43, 136.08, 135.73,
130.17, 129.74, 129.61, 129.42, 129.31, 129.03, 128.56, 113.94,
112.96, 106.74, 105.01, 55.19, 53.23, 46.67, 15.13, 3.56, 2.97; IR
(neat) 3328, 3005, 2831,1714, 1655, 1590, 1567, 1472, 1363, 1223,
1026, 813, 698 cmꢀ 1; LRMS (ESI) m/z calculated for C31H28N8O2 [M +
H]+ 545.23, found 544.99.
C
24H20F3N7O [M + H]+ 480.18, found 480.16; HPLC purity 100% ee.
4.2.3.12. (S)-2-(((7H-Purin-6-yl)amino)(cyclopropyl)methyl)-5-methyl-
3-phenylquinazolin-4(3H)-one (6m). Synthesized from general proced-
ure C for 15 h using compound 5m. 62% yield of the product as a white
solid; mp 183–185 C; [α D20
]
+ 93.3759(c 0.5, CH2Cl2); 1H NMR (400
◦
MHz, CDCl3) δ: 13.90 (brs, 1H), 8.31 (s, 1H), 7.97 (s, 1H), 7.61–7.51 (m,
5H), 7.41 (d, J = 8 Hz, 1H), 7.21–7.19 (m, 1H), 7.00 (d, J = 8 Hz, 1H),
4.94 (m, 1H), 2.81 (s, 3H), 1.42–1.37 (m, 1H), 0.49–0.40 (m, 3H),
0.25–0.23 (m, 1H); 13C NMR (100 MHz, CDCl3) δ: 162.97, 156.13,
153.67, 151.93, 149.68, 148.77, 141.52, 138.42, 136.48, 133.65,
129.75, 129.63, 129.61, 129.36, 128.98, 125.61, 119.43, 53.38, 23.05,
15.23, 3.58, 2.98; IR (neat) 3380, 3198, 3008, 2812, 1738, 1680, 1588,
1470, 1288, 1228, 1131, 1043, 810, 749, 697 cmꢀ 1; LRMS (ESI) m/z
calculated for C24H21N7O [M + H]+ 424.18, found 423.94; HPLC purity
100% ee.
4.2.5. General procedure E for the preparation of compound 8a-c
Trifluoroacetic acid (0.15 M) was added to a stirred solution of the
◦
corresponding compound (7–9, 1 eq.) in CH2Cl2 (0.3 M) at 0 C. The
resulting reaction mixture was stirred at room temperature for 2 h. The
completion of the reaction was monitored by TLC on silica (CH2Cl2-
MeOH = 10:1). The reaction mixture was quenched with 1 M NaOH (pH
~ 7) at 0 ◦C, and then the aqueous layer was extracted with CH2Cl2. The
combined extracts were dried over MgSO4, filtered, and evaporated
under reduced pressure. The crude residue was subjected to flash silica
gel column chromatography (CH2Cl2-MeOH gradient or EtOAc-MeOH
gradient) to afford title compounds.
4.2.4. General procedure d for the preparation of compound 7a-c
In a sealed tube, 4-methoxy benzylamine (1 eq.) was added to a
stirred solution of the appropriate compound (6b, 6c, and 6f) (1 eq.) and
TEA (1.44 M) dissolved in ethanol (0.1 M). The reaction mixture was
heated to 160–180 ◦C for 3–4 days. The completion of the reaction was
monitored by TLC on silica (CH2Cl2-MeOH = 10:1). After cooling to
room temperature, the reaction mixture was concentrated under
reduced pressure. The crude residue was subjected to flash silica gel
column chromatography (CH2Cl2-MeOH gradient) to afford title
compounds.
4.2.5.1. (S)-2-(1-((7H-Purin-6-yl)amino)ethyl)-5-amino-3-phenyl-
quinazolin-4(3H)-one (8a). Synthesized from general procedure E using
compound 7a. 77% yield of the product as an ivory solid; mp 161 ◦C;
[
α 2D0
]
+ 138.8879(c 0.5, CH2Cl2); 1H NMR (400 MHz, CD3OD) δ: 8.19 (s,
1H), 8.11 (s, 1H), 7.61–7.56 (m, 2H), 7.49–7.44 (m, 3H), 7.40 (t, J = 8
Hz, 1H), 6.80 (dd, J = 8 Hz, 0.8 Hz, 1H), 6.63 (dd, J = 8 Hz, 0.8 Hz, 1H),
6.16 (s, 2H), 5.00–4.98 (m, 1H), 1.51 (d, J = 6.8 Hz, 3H); 13C NMR (100
MHz, MeOD) δ: 164.51, 157.01, 152.18, 151.12, 148.50, 139.63,
136.26, 135.02, 129.28, 129.27, 128.97, 128.90, 112.61, 111.46,
104.66, 18.35; IR (neat) 3344, 2922, 2851, 1663, 1604, 1576, 1463,
1245, 1137, 1017, 816, 755, 699 cmꢀ 1; LRMS (ESI) m/z calculated for
C21H18N8O [M + H]+ 399.16, found 399.17; HPLC purity 91.83% ee.
4.2.4.1. (S)-2-(1-((7H-purin-6-yl)amino)ethyl)-5-((4-methoxybenzyl)
amino)-3-phenylquinazolin-4(3H)-one (7a). Synthesized from general
procedure D using the compound 6b. 43% yield of the product as pale
yellow solid; mp 130 ◦C in decomposition; [α 2D0
+ 204.8238(c 0.5,
]
CH2Cl2); 1H NMR (400 MHz, CDCl3) δ: 8.81–8.87 (m, 1H), 8.32 (s, 1H),
7.95 (s, 1H), 7.59–7.49 (m, 5H), 7.36–7.34 (m, 1H), 7.25–7.23 (m, 2H),
7.13 (m, 1H), 6.87 (d, J = 7.6 Hz, 1H), 6.81 (d, J = 8.8 Hz, 2H), 6.47 (d,
J = 8.4 Hz, 1H), 5.17 (m, 1H), 4.32–4.30 (m, 2H), 3.75 (s, 3H), 1.48 (d, J
= 6.8 Hz, 3H); 13C NMR (100 MHz, CDCl3) δ: 164.68, 158.78, 156.75,
153.44, 152.06, 150.48, 148.91, 138.38, 136.05, 135.82, 130.22,
130.00, 129.96, 129.51, 129.00, 128.91, 128.61, 114.02, 112.94,
106.87, 105.06, 55.25, 46.73, 29.72, 20.31; IR (neat) 3318, 3010, 2950,
2882, 1780, 1655, 1566, 1509, 1410, 1328, 1247, 1143, 1033, 908, 813,
698 cmꢀ 1; LRMS (ESI) m/z calculated for C29H26N8O2 [M + H]+ 519.22,
found 519.29.
4.2.5.2. (S)-2-(1-((7H-Purin-6-yl)amino)propyl)-5-amino-3-phenyl-
quinazolin-4(3H)-one (8b). Synthesized from general procedure E using
◦
compound 7b. 73% yield of the product as yellow solid; mp 174 C;
1
[
α 2D0
]
+ 185.7359(c 0.5, CH2Cl2); H NMR (400 MHz, CDCl3) δ: 13.79
(brs, 1H), 8.31 (s, 1H), 7.96 (s, 1H), 7.61–7.34 (m, 6H), 7.03 (d, J = 8
Hz, 1H), 6.88 (d, J = 8 Hz, 1H), 6.51 (d, J = 8 Hz, 1H), 6.18 (s, 2H), 5.13
(m, 1H), 2.04–1.93 (m, 1H), 1.83–1.75 (m, 1H), 0.85 (t, J = 7.2 Hz, 3H);
13C NMR (100 MHz, CDCl3) δ: 164.47, 155.91, 154.10, 152.03, 150.18,
148.52, 138.35, 136.05, 135.29, 129.94, 129.54, 129.44, 129.03,
114.31, 111.82, 52.24, 27.57, 10.06; IR (neat) 3343, 3040, 2818, 1714,
1660, 1574, 1470, 1293, 1131, 1013, 816, 751, 698 cmꢀ 1; LRMS (ESI)
4.2.4.2. (S)-2-(1-((7H-purin-6-yl)amino)propyl)-5-((4-methoxybenzyl)
amino)-3-phenylquinazolin-4(3H)-one (7b). Synthesized from general
12

Y.S. Kim et al.
Bioorganic & Medicinal Chemistry 45 (2021) 116312
m/z calculated for C22H20N8O [M + H]+ 413.18, found 413.33; HPLC
14 Crabbe T, Welham MJ, Ward SG. The PI3K inhibitor arsenal: choose your weapon.
Trends Biochem Sci. 2007;32:450–456.
purity 98.42% ee.
15 Dreyling M, Santoro A, Mollica L, et al. Phosphatidylinositol 3-Kinase Inhibition by
Copanlisib in Relapsed or Refractory Indolent Lymphoma. J Clin Oncol. 2017;35:
3898–3905.
4.2.5.3. (S)-2-(((7H-Purin-6-yl)amino)(cyclopropyl)methyl)-5-amino-3-
phenylquinazolin-4(3H)-one (8c). Synthesized from general procedure E
using compound 7c. 60% yield of the product as a white solid; mp
16 Pirali T, Ciraolo E, Aprile S, et al. Identification of a potent phosphoinositide 3-kinase
pan inhibitor displaying a strategic carboxylic acid group and development of its
prodrugs. Chem Med Chem. 2017;12:1542–1554.
173.5 ◦C; [α 1D6.5
]
+ 184.378(c 0.5, CH2Cl2); 1H NMR (400 MHz, CDCl3) δ:
17 Ma CC, Zhang CM, Tang LQ, Liu ZP. Discovery of novel quinazolinone derivatives as
high potent and selective PI3Kδ and PI3Kδ/γ inhibitors. Eur J Med Chem. 2018;151:
9–17.
13.80 (brs, 1H), 8.30 (s, 1H), 7.97 (s, 1H), 7.61–7.38 (m, 6H), 6.95–6.90
(m, 2H), 6.53 (d, J = 8 Hz, 1H), 6.16 (s, 2H), 4.91 (m, 1H), 1.40–1.35 (m,
1H), 0.49–0.39 (m, 3H), 0.24–0.21 (m, 1H); 13C NMR (100 MHz, CDCl3)
δ: 164.44, 155.60, 153.56, 151.88, 150.16, 148.54, 138.44, 136.08,
135.23, 129.70, 129.62, 129.38, 129.29, 129.05, 114.18, 111.76,
105.46, 53.23, 15.14, 3.56, 2.95; IR (neat) 3439, 3340, 3002, 1659,
1601, 1673, 1470, 1292, 1234, 1133, 1071, 1023, 935, 816, 749, 698
cmꢀ 1; LRMS (ESI) m/z calculated for C23H20N8O [M + H]+ 425.18,
found 425.15; HPLC purity 100% ee.
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Declaration of Competing Interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
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Acknowledgments
This research was supported by a grant of the Korea Health Tech-
nology R&D Project through the Korea Health Industry Development
Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic
of Korea (grant number: HI18C1942)
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