Extensive investigation of benzylic N-containing substituents on the pyrrolopyrimidine skeleton as Akt inhibitors with potent anticancer activity

Article abstract of DOI:10.1016/j.bioorg.2020.103671

Continuous optimization of benzylic substituents on 1-(4-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-2-phenylethan-1-one structure as Akt inhibitors was described in this paper. Particularly, compounds 8 and 14g exhibited high enzymatic potency against all Akt isoforms and antiproliferative effects in mantle cell lymphoma cell lines, as well as favorable cytotoxicities in patient primary cancer cells. Low micromolar doses of both 8 and 14g dose-dependently induced cell apoptosis and G₂/M cell cycle arrest, also suppressed the phosphorylation level of Akt downstream targets GSK3β and S6.

Full text of DOI:10.1016/j.bioorg.2020.103671

Journal Pre-proofs
Extensive Investigation of Benzylic N-containing Substituents on the Pyrrolo-
pyrimidine Skeleton as Akt Inhibitors with Potent Anticancer Activity
Yang Liu, Zhen Zhang, Fansheng Ran, Kaiwen Guo, Xin Chen, Guisen Zhao
PII:
S0045-2068(19)32162-5
DOI:
https://doi.org/10.1016/j.bioorg.2020.103671
YBIOO 103671
Reference:
To appear in:
Bioorganic Chemistry
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Revised Date:
Accepted Date:
18 December 2019
7 February 2020
14 February 2020
Please cite this article as: Y. Liu, Z. Zhang, F. Ran, K. Guo, X. Chen, G. Zhao, Extensive Investigation of
Benzylic N-containing Substituents on the Pyrrolopyrimidine Skeleton as Akt Inhibitors with Potent Anticancer
Activity, Bioorganic Chemistry (2020), doi: https://doi.org/10.1016/j.bioorg.2020.103671
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Extensive Investigation of Benzylic N-containing Substituents on the
Pyrrolopyrimidine Skeleton as Akt Inhibitors with Potent Anticancer Activity
Yang Liu1, 2, Zhen Zhang1, Fansheng Ran1, Kaiwen Guo1, Xin Chen1 and Guisen Zhao1, *
1Department of Medicinal Chemistry, Key Laboratory of Chemical Biology (Ministry of Education),
School of Pharmaceutical Sciences, Shandong University, Jinan, Shandong 250012, PR China;
2Department of Lymphoma & Myeloma, The University of Texas MD Anderson Cancer Center, Houston,
TX 77030, USA.
Corresponding author
Guisen Zhao*
Department of Medicinal Chemistry, School of Pharmaceutical Sciences, Shandong University, Jinan,
Shandong 250012, PR China;
Tel/ Fax: +86 531 88382009;
E-mail: guisenzhao@sdu.edu.cn.

Abstract
Continuous optimization of benzylic substituents on 1-(4-(7H-pyrrolo[2,3-d]pyrimidin-4-
yl)piperazin-1-yl)-2-phenylethan-1-one structure as Akt inhibitors was described in this paper.
Particularly, compounds 8 and 14g exhibited high enzymatic potency against all Akt isoforms and
antiproliferative effects in mantle cell lymphoma cell lines, as well as favorable cytotoxicities in patient
primary cancer cells. Low micromolar doses of both 8 and 14g dose-dependently induced cell apoptosis
and G2/M cell cycle arrest, also suppressed the phosphorylation level of Akt downstream targets GSK3β
and S6.
Keywords: Akt, Anticancer, Docking, Mantle cell lymphoma, Pyrrolopyrimidines
1. Introduction
PI3K/Akt signaling pathway is one of the most frequently activated pathways in human cancers,
targeted agents against this pathway represent effective anticancer therapies [1]. Akt, a critical mediator
in PI3K/Akt pathway, is an evolutionarily conserved serine/threonine kinase with three closely related
isoforms, Akt1, Akt2 and Akt3, all of which share highly conserved domains: a N-terminal PH domain,
a central kinase domain and a C-terminal regulatory domain [2]. Akt modulates cell survival and
proliferation through pleiotropic downstream substrates such as glycogen synthase kinase 3 (GSK-3) and
mammalian target of rapamycin complex 1 (mTORC1). Overexpression of Akt is a common feature
identified in hematological neoplasms and solid tumors [3-5], and often associated with resistance to
chemotherapy or radiotherapy [6]. Taken together, these factors would highly recommend Akt as a
compelling target in cancer treatment [7].
Mantle cell lymphoma (MCL) is an uncommon subtype of B-cell non-Hodgkin lymphoma with
rapid clinical evolution and unfavorable outcomes [8, 9]. Understanding the molecular pathophysiology
for MCL has resulted in the identification of innovative targeted therapeutic agents [10]. Constitutive
activation of Akt has been validated to contribution of pathogenesis and survival for mantle cell
lymphoma [11], and considerable research efforts have validated that Akt inhibitors effectively decreased
MCL tumor cell growth, suggesting Akt inhibition represents an innovative therapeutic strategy for MCL
[12, 13].
Various attempts on structural optimization of pyrrolopyrimidine based ATP competitive inhibitors
have been reported in our previous studies [14-16]. Inspired by the successful introduction of the rigid
piperazine moiety, the benzylic position was taken as a continuous optimization site, with various

aliphatic amino groups and nitro-containing heterocycles introduced to explore further activity
improvement on the pyrrolopyrimidine scaffold.
Fig. 1. Optimization schema for the designed compounds.
2. Results and discussion
2.1. Chemistry
The general synthetic route for the pyrrolopyrimidine derivatives is outlined in Scheme 1.
Bromination on the 5-position of the commercially available 1 afforded compound 2, subsequent
introduction of the tert-butoxycarbonyl (Boc) protected piperazine linker via SNAr delivered 3, followed
by removal of the Boc group affording 4 as dihydrochloride salts. Boc-protection of intermediate 5 and
further amide coupling accomplished the first target compound 7. Removal of the Boc moiety provided
resultant 8 as hydrochloride salt.
Commercially available 9 reacted with methanol under H2SO4 condition to give 10, followed by
free radical reaction at the benzylic position produced 11, which were further alkylated to give substituted
amines 12a-12h, subsequent ester hydrolysis gave the carboxylic acids 13a-13h. Amide coupling
between 13a-13h and intermediate 4 yielded compounds 14a-14h. Final deprotection of the Boc group
accomplished
target
compounds
15a-15h.

Scheme 1. Synthetic route for the designed derivatives. Reagents and conditions: (a) NBS, DMF, r.t.; (b) 1-Boc-piperazine, DIEA,
DMF, 110°C; (c) HCl-dioxane, CH3OH, r.t.; (d) (Boc)2O, NaOH (aq), THF, r.t.; (e) EDCI, HOBt, DIEA, DMF, r.t.; (f) HCl-
dioxane, CH3OH, r.t.; (g) CH3OH, H2SO4, 60°C; (h) NBS, AIBN, CCl4, reflux; (i) Amines, DIEA, CH3CN, 80°C; (j) CH3CH2OH,
NaOH (aq, 10M), 80°C; (k) EDCI, HOBt, DIEA, DMF, r.t.; (l) HCl-dioxane, CH3OH, r.t..
2.2. Akt inhibition and cell antiproliferative activity
The Akt1 inhibitory activities of the synthesized pyrrolopyrimidine compounds were determined
via a Homogeneous Time-Resolved Fluorescence (HTRF) kinase activity assay. GSK690693, first Akt
inhibitor entered clinical trials, was used as the positive control.

Table 1. The inhibitory effects of pyrrolopyrimidine derivatives on Akt1 activity
Akt1 inhibition (%) Akt1 inhibition (%)
Code
7
R
Code
R
1 μM
50 nM
1 μM
50 nM
O
N
Boc
HN
58.4
10.4
14e
65.6
22.8
Boc
N
8
83.3
88.5
83.1
73.0
14f
29.6
84.1
NAa
80.8
2
N
N
N
N
14a
14g
Boc
HN
14b
14c
14d
81.8
95.5
78.0
52.6
14h
15a
15b
60.6
93.7
93.8
21.9
72.7
55.1
N
N
N
N
H
N
N
NH2
N
41.1
98.4
38.8
83.6
GSK690693
aNA means not active
The structure-activity-relationship of the substituents at the benzylic position is shown in Table 1.
The introduction of the free amino analogue 8 was proved to be the most potent resultant compound
(83.1% inhibition at 50 nM), which showed comparable inhibitory activity to that of GSK690693.
Dimethylamino and diethylamino substituents (14a and 14b) decreased Akt1 inhibitory activities to a
light degree, while most of the nitrogen-containing heterocyclic substitutions remarkably decreased the
enzyme potency, except for methylpiperazine 14g and piperazine 15a (80.8% and 72.7% inhibition at 50
nM, respectively). The bulky Boc groups (7, 14f and 14h) were not well tolerated at the benzylic position,
which resulted in a substantial loss of enzyme potency.
Compounds with inhibition rate higher than 70% at 50 nM were further tested their
antiproliferative effects against multiple MCL cell lines. Ibrutinib, a covalent BTK inhibitor approved

by FDA to treat MCL, was employed as positive control with GSK690693. In comparison with
GSK690693 and ibrutinib, all tested compounds displayed improved antiproliferative activities in
MCL cell lines. As anticipated, compounds 8 and 14g with preferable Akt1 enzyme potency also
displayed favorable antiproliferative activities with IC50 values in low micromolar range.
Table 2. Cell proliferation assay assessing the effects of 8 and 14g on MCL cell lines
Cell viability assay, IC50 ± SD/μMa
Code
Granta-519
10.5 ± 1.4
18.5 ± 1.6
1.3 ± 0.6
1.6 ± 0.5
2.6 ± 0.3
0.9 ± 0.3
2.5 ± 0.7
Mino
Jeko-1
Maver-1
Z-138
Rec-1
Ibrutinib
6.1 ± 1.4 2.5 ± 0.2 2.7 ± 1.2 2.8 ± 0.4
5.6 ± 0.4
GSK690693
6.4 ± 0.2 3.3 ± 2.6 3.6 ± 2.1 5.1 ± 0.5 14.9 ± 1.7
8
1.6 ± 0.1 1.3 ± 0.4 1.4 ± 0.2 1.1 ± 0.2
1.6 ± 0.3 1.1 ± 0.1 1.9 ± 0.1 0.7 ± 0.2
1.9 ± 0.8 1.9 ± 0.1 2.1 ± 0.3 1.6 ± 0.7
1.1 ± 0.2 0.8 ± 0.1 0.7 ± 0.2 0.7 ± 0.1
1.6 ± 0.6
0.9 ± 0.2
1.5 ± 0.4
0.6 ± 0.1
14a
14b
14g
15a
2.3 ± 0.4 1.4 ± 0.4 1.2 ± 0.6 1.4 ± 0.4 25.5 ± 2.7
aValues are means of three independent experiments
2.3 Akt isoforms selectivity assay
The FRET based Z'-LYTE kinase assay was used to determine the capability of compounds 8 and
14g to inhibit all three Akt isoforms. The selectivity profile was summarized in Table 3, compounds 8
and 14g were both validated as pan-Akt inhibitors with IC50 values in the low nanomolar range, and the
IC50 values for Akt2 were within 3-7 folds of Akt1 and Akt3.
Table 3. Akt isoforms inhibitory profile of 8 and 14g
Akt kinase assay (IC50, nM)
Code
Akt1
3.2
Akt2
20.9
20.2
Akt3
6.2
8
14g
3.4
4.7
2.4. Cell apoptosis and cell cycle assay
To identify the mechanisms underlying the antiproliferative effects of these inhibitors, the efforts
were focused on possible impact on cell apoptosis and cell cycle progression, which is mediated by Akt

downstream factors [17]. Accordingly, Annexin V/PI apoptosis and PI cell cycle assays were
performed after 8 and 14g treatment in Jeko-1 and Mino cells. Fig. 2 and Supplementary Fig. 1 show
the dose dependent apoptosis after exposure of Jeko-1 and Mino cells to 8 or 14g for 24 hours,
suppression of the Akt downstream death promoter Bad was also validated by western blotting, as well
as activated caspase-3 and cleaved PARP (Fig. 3A). Cell cycle assay demonstrated that 8 or 14g
treatment caused a significant G2/M phase arrest in both Jeko-1 and Mino cells.
Fig. 2. Cell apoptosis (A) and cell cycle assay (B) in Jeko-1 cells after 24 hours treatment of 8 and 14g at indicated
concentrations. The data shown are the mean of three independent experiments. Statistical significance is indicated as *P < 0.05,
**P < 0.01 compared to DMSO control.
2.5. Inhibitory effects on Akt signaling
Western blot analysis was performed to detect the capability of 8 and 14g on the phosphorylation
of Akt and its downstream effectors in Jeko-1 cells, in which PTEN loss has been validated as the
genetic mutation that drives constitutively active PI3K/AKT signaling [18]. After 24 hours of
treatment, both 8 and 14g dose-dependently downregulated phosphorylated GSK3β and S6, which are
major downstream targets of Akt (Fig. 3A). Hyperphosphorylation of the two Akt regulatory sites
(Ser473 and Thr308) was also confirmed, which is consistent with the observations from certain

reported Akt inhibitors, proposing a possible positive feedback loop involved [19-21].
2.6. Cell viability assay in primary MCL cells
Cell viability assay was further performed for 8 and 14g against the tumor cells isolated from two
MCL patients (PT1 and PT2, Supplemental Fig. 2). As shown in Fig. 3B, 8 and 14g dramatically
decreased the cell viability of the two tested MCL patient cells compared to that of GSK690693 and
ibrutinib. Subsequent western blot validation on the primary cells was also in agreement with that
observed in Jeko-1 cells (Fig. 3C).
Fig. 3. Western blot analysis in Jeko-1 cells and efficacy validation in patient primary cancer cells. (A) Effects on apoptosis
signaling and Akt signaling in Jeko-1 cells after 24 hours treatment of 8 and 14g; (B) Cell viability assay in two primary MCL
cells. Values were normalized to DMSO control; (C) Effect validation on Akt signaling of 8 and 14g in PT1 primary cells.
2.7. Molecular modeling
The interaction modes between Akt1 and candidates 8 and 14g were proposed and simulated by
Sybyl 2.0, key interactions formed were highlighted and depicted in Fig. 4. Both inhibitors formed
bidentate hydrogen bonds with residues Ala230 and Glu228 in the hinge region. The 4-chlorophenyl
group projected into a hydrophobic pocket under the P-loop for hydrophobic contact. A beneficial
hydrogen bond was formed between Glu234 and the primary amine of 8, while the methylpiperazine
moiety of 14g interacted with residue Arg4 via two hydrogen bonds, which enhanced the favorable
binding affinity for the resultant chimeric conformation.

Fig. 4. Proposed binding modes of 8 (left) and 14g (right) at the ATP binding site of Akt1 (PDB: 3MV5).
3. Conclusion
In summary, modification on the benzylic position of pyrrolopyrimidine core was extensively
carried out. The promising pan-Akt inhibitors 8 and 14g sustained the in vitro Akt1 inhibition and
cellular inhibitory activity against multiple cancer cells, as well as desirable effects in patient primary
cells, endorsing further development of the two candidates as anticancer agents.
4. Experimental section
4.1. Chemistry
All solvents and reagents were purchased from commercially available sources and used without
further purification. Reactions were monitored by thin layer chromatography (TLC), and silica gel GF254
plates were used and visualized with UV light. Column chromatography was performed with silica gel
using the solvents indicated. NMR spectra were recorded on a Bruker avance DRX600 or 400
Spectrometer using TMS as an internal standard in DMSO-d6. Chemical shifts were reported in parts per
million (ppm). Coupling constants (J) were given in Hz. The mass spectra (MS) were measured with an
API 4000. All of the melting points were determined in a Büchi capillary melting point apparatus and
are uncorrected. Human Akt1 was obtained from Carna Biosciences, Inc. (Canada). The HTRF assay kit
was purchased from Cisbio assays, Inc. (France).
4.2. General synthesis of compounds
4.2.1. General synthesis of compound 4
The synthesis of 4 is the same as previously descried in our paper [14].
4.2.2. Tert-butyl bis(2-chloroethyl)carbamate (6)
To a solution of 5 (5 mmol) in tetrahydrofuran (20 mL), 10% sodium hydroxide solution (6 mmol)

was added. This was followed by addition of di-tert-butyl dicarbonate (7.5 mmol) over a period of 10
min at 0℃. Reaction mixture was stirred for 3 hours at room temperature after the addition, and acidified
to pH 2 with 1N hydrochloric acid, the product thus obtained was extracted with ethyl acetate (3×50
mL). The organic layer was combined, washed with brine, dried over sodium sulfate and evaporated
under reduced pressure to give the product 6 as a clear oil, yield 71.3%. Compound 6 was used directly
for the next step without further purification.
4.2.3. General synthesis of compound 7
To a solution of 6 (1.75 mmol) and N, N-Diisopropylethylamine (DIEA, 8.75 mmol) in
dimethylformamide (DMF, 2 mL) was added 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide
hydrochloride (EDCI) (2.1 mmol) and 1-hydroxybenzotriazole (HOBt) (2.1 mmoL). The reaction
mixture was stirred at 0℃ for 1 hour, followed by addition of 4 (1.75 mmol). After completion of addition,
the final mixture was stirred at room temperature for another 24 hours, then poured into ice water (20
mL), washed with saturated aqueous sodium bicarbonate (3×20 mL) and brine (3×20 mL), dried over
sodium sulfate and solvent was evaporated under reduced pressure. The crude product was purified by
silica gel chromatography eluting with dichloromethane /methanol (50:1) to give 7. Brown solid, yield
43.8%, mp: 128~131℃, 1H NMR (400 MHz, MeOD) δ (ppm): 8.24 (s, 1H), 7.39 (s, 4H), 7.34 (s, 1H),
5.67 (s, 1H), 3.95-3.84 (m, 1H), 3.84-3.72 (m, 2H), 3.72-3.64 (m, 1H), 3.64-3.53 (m, 2H), 3.53-3.42 (m,
1H), 3.31 (dt, J = 3.2, 1.6 Hz, 3H), 3.21-3.10 (m, 1H), 1.43 (s, 9H). 13C NMR (100 MHz, MeOD) δ
(ppm): 169.20, 159.69, 155.87, 151.33, 150.39, 136.03, 133.96, 129.35, 128.79, 123.95, 105.14, 86.58,
79.51, 54.66, 49.76, 49.50, 45.14, 42.12, 27.28. MS (ESI) m/z: 549 [M+H]+.
4.2.4. General synthesis of compounds 8
7 (2 mmol) was suspended in methanol (2 mL), after addition of the 4M HCl in dioxane (3 ml), the
reaction mixture was stirred for 48 hours at room temperature. The precipitate was filtrated, the obtained
solid was recrystallized from methanol to give the desired 8. Brown solid, yield: 72.5%, mp: 264~266℃,
1H NMR (400 MHz, D2O) δ (ppm): 8.08 (s, 1H), 7.39 (d, J = 8.6 Hz, 2H), 7.35 (d, J = 8.7 Hz, 2H), 7.32
(s, 1H), 5.50 (s, 1H), 3.99-3.88 (m, 1H), 3.86-3.73 (m, 1H), 3.71-3.47 (m, 5H), 3.46-3.34 (m, 1H), 3.02-
2.90 (m, 1H). 13C NMR (100 MHz, D2O) δ (ppm): 166.38, 154.28, 146.67, 143.18, 136.26, 130.14,
129.94, 129.51, 126.60, 103.01, 88.85, 66.52, 54.53, 49.63, 48.75, 44.33, 41.97. MS (ESI) m/z: 449
[M+H]+.
4.2.5. General synthesis of compounds 10

A solution of 9 (40 mmol) in dry methanol (25 mL) was treated with 5 drops of concentrated
H2SO4 (cat.) at room temperature. The reaction resulting mixture was stirred and heated at reflux for 5
hours to completion and was concentrated under reduced pressure. The crude product was dissolved in
25 mL ethyl acetate, washed with saturated aqueous sodium bicarbonate, water (3×25mL) and brine
(3×25mL), dried over Na2SO4, filtered and concentrated to get the titled compound 10 as a yellow oil,
yield 97.6%, 1H NMR (400 MHz, DMSO-d6) δ (ppm): 7.39 (d, J = 8.4 Hz, 2H), 7.30 (d, J = 8.4 Hz, 2H),
3.70 (d, 2H), 3.61 (s, 3H). MS (ESI) m/z: 185 [M+H]+.
4.2.6. General synthesis of compounds 11
To a solution of ethyl 4-pyridylacetate (27.95 mmol) in carbon tetrachloride (20 mL), N-
bromosuccinimide (27.95 mmol) and Azobisisobutyronitrile (AIBN, 7.95 mmol) were added. The
reaction mixture was heated at reflux for 2 hours after which the solids were removed by filtration. The
organics filtrate was concentrated under reduced pressure. The resulting crude oil was purified by flash
chromatography on silica gel to obtain as a yellow oil, 94.3%, 1H NMR (400 MHz, DMSO-d6) δ (ppm):
7.59 (d, J = 8.4 Hz, 2H), 7.49 (d, J = 8.4 Hz, 2H), 6.00 (s, 1H), 3.73 (s, 3H). MS (ESI) m/z: 263 [M+H]+.
4.2.7. General synthesis of compounds 12
To a solution of compound 11 (2 mmol) in acetonitrile (5 mL) was added substituted amine (2.2
mmol) and DIEA (3 mmol) at room temperature. The reaction mixture was refluxed for 4 hours and
then the solvent was evaporated in vacuo. The resultant residue was diluted with ethyl acetate (20 mL),
washed with water and brine (3×20 mL), dried over Na2SO4.The mixture was then filtered and the
filtrate was concentrated in vacuo. The resulting residue was purified by flash column chromatography
with ether/ethylacetate (1/3) to afford 12.
4.2.7.1. Methyl 2-(4-chlorophenyl)-2-(dimethylamino)acetate (12a)
Yellow oil, yield 61.8%, 1H NMR (400 MHz, CDCl3) δ (ppm): 7.39 (d, J = 8.8 Hz, 2H), 7.33 (d, J
= 8.8 Hz, 2H), 3.86 (s, 1H), 3.70 (s, 3H), 2.24 (s, 6H). MS (ESI) m/z: 228 [M+H]+.
4.2.7.2. Methyl 2-(4-chlorophenyl)-2-(diethylamino)acetate (12b)
Yellow oil, yield 70.2%, 1H NMR (400 MHz, CDCl3) δ (ppm): 7.39 (d, J = 8.4 Hz, 2H), 7.31 (d, J
= 8.8 Hz, 2H), 3.94 (s, 1H), 3.68 (s, 3H), 2.36 (q, 4H), 1.60 (t, 6H). MS (ESI) m/z: 256 [M+H]+.
4.2.7.3. Methyl 2-(4-chlorophenyl)-2-(pyrrolidin-1-yl)acetate (12c)
Colorless oil, yield 90%, 1H NMR (400 MHz, CDCl3) δ (ppm): 7.43 (d, J = 8.4 Hz, 2H), 7.32 (d,
J = 8.4 Hz, 2H), 3.90 (s, 1H), 3.68 (s, 3H), 2.55-2.43 (m, 4H), 1.82-1.81 (m, 4H). MS (ESI) m/z: 254

[M+H]+.
4.2.7.4. Methyl 2-(4-chlorophenyl)-2-(3,5-dimethylpiperidin-1-yl)acetate (12d)
Colorless oil, yield 65.7%, 1H NMR (400 MHz, CDCl3) δ (ppm): 7.31-7.26 (m, 4H), 4.45 (s, 1H),
3.69 (s, 3H), 2.73-2.67 (m, 2H), 2.70-1.84 (m, 6H), 1.07 (t, J = 6.0 Hz, 6H). MS (ESI) m/z: 296 [M+H]+.
4.2.7.5. Methyl 2-(4-chlorophenyl)-2-morpholinoacetate (12e)
Clear oil, yield 71.4%, 1H NMR (400 MHz, DMSO-d6) δ (ppm): 7.45 (d, J = 9.2 Hz, 2H), 7.43 (d,
J = 9.2 Hz, 2H), 4.16 (s, 1H), 3.61 (s, 3H), 3.57 (d, J = 4.4 Hz, 4H), 2.35 (d, J = 4.4 Hz, 4H). MS (ESI)
m/z: 270 [M+H]+.
4.2.7.6. Tert-butyl 4-(1-(4-chlorophenyl)-2-methoxy-2-oxoethyl)piperazine-1-carboxylate (12f)
Clear oil, yield 99.5%, 1H NMR (400 MHz, CDCl3) δ (ppm): 7.38 (d, J = 8.4 Hz, 2H), 7.33 (d, J =
8.4 Hz, 2H), 3.99 (s, 1H), 3.69 (s, 3H), 3.44 (s, 4H), 2.39 (s, 4H), 1.44 (s, 9H). MS (ESI) m/z: 369 [M+H]+.
4.2.7.7. Methyl 2-(4-chlorophenyl)-2-(4-methylpiperazin-1-yl)acetate (12g)
Brown oil, yield 66.7%, 1H NMR (400 MHz, CDCl3) δ (ppm): 7.39 (d, J = 8.4 Hz, 2H), 7.32 (d, J
= 8.4 Hz, 2H), 3.96 (s, 1H), 3.68 (s, 3H), 2.63-2.40 (m, 8H), 2.31 (s, 3H). MS (ESI) m/z: 283 [M+H]+.
4.2.7.8. Methyl 2-(4-((tert-butoxycarbonyl)amino)piperidin-1-yl)-2-(4-chlorophenyl)acetate (12h)
White solid, yield 70.6%, 1H NMR (400 MHz, CDCl3) δ (ppm): 7.37 (d, J = 8.4 Hz, 2H), 7.32 (d,
J = 8.4 Hz, 2H), 4.43 (br, 1H), 3.96 (s, 1H), 3.68 (s, 3H), 3.49 (br, 1H), 2.86 (br, 1H), 2.67 (br, 1H), 2.24
(t, J = 10.8 Hz, 1H), 2.01-1.84 (m, 3H), 1.45-1.43 (s, 10H). MS (ESI) m/z: 383 [M+H]+.
4.2.8. General synthesis of compounds 13
To a solution of 12 (1 mmol) in 3 mL of ethanol/water (2/1), 2mL of 1 N aqueous sodium hydroxide
solution (2 mmol) were added and the mixture was stirred at room temperature for 4 hours. The organic
solvents were removed under reduced pressure and the resulting aqueous solution was adjusted in an ice
bath to pH 6 using 1 N aqueous hydrochloric acid. The resulting solid was filtered, washed with water to
obtain 13.
4.2.8.1. 2-(4-Chlorophenyl)-2-(dimethylamino)acetic acid (13a)
White solid, yield 81.5%, 1H NMR (400 MHz, CDCl3) δ (ppm): 7.41 (d, J = 7.2 Hz, 2H), 7.16 (d,
J = 7.2 Hz, 2H), 4.77 (s, 1H), 2.84 (s, 6H). MS (ESI) m/z: 212 [M-H]-.
4.2.8.2. 2-(4-Chlorophenyl)-2-(diethylamino)acetic acid (13b)
White solid, yield 20%, 1H NMR (400 MHz, MeOD) δ (ppm): 7.64-7.54 (m, 2H), 7.48-7.41 (m,
2H), 4.56 (s, 1H), 3.21 (td, J = 13.3, 6.4 Hz, 2H), 3.05 (s, 2H), 1.25 (t, J = 7.1 Hz, 6H). MS (ESI) m/z:

240 [M-H]-.
4.2.8.3. 2-(4-Chlorophenyl)-2-(pyrrolidin-1-yl)acetic acid (13c)
White solid, yield 86.9%, 1H NMR (400 MHz, CD3OD) δ (ppm): 7.56 (d, J = 8.8 Hz, 2H), 7.46
(d, J = 8.4 Hz, 2H), 4.51 (s, 1H), 3.31-3.29 (m, 4H), 2.11-1.99 (m, 4H). MS (ESI) m/z: 238 [M-H]-.
4.2.8.4. 2-(4-Chlorophenyl)-2-(3,5-dimethylpiperidin-1-yl)acetic acid (13d)
White solid, yield 58.8%, 1H NMR (400 MHz, DMSO-d6) δ (ppm): 7.43 (d, J = 8.8 Hz, 2H), 7.35
(d, J = 8.8 Hz, 2H), 3.81 (s, 1H), 3.09-3.07 (m, 1H), 2.56-2.54 (m, 1H), 1.70-1.63 (m, 4H), 1.51 (t, J =
10.8 Hz, 1H), 0.80 (d, J = 5.2 Hz, 3H), 0.70 (d, J = 8.4 Hz, 3H), 0.54-0.45 (m, 1H). MS (ESI) m/z: 280
[M-H]-.
4.2.8.5. 2-(4-Chlorophenyl)-2-morpholinoacetic acid (13e)
White solid, yield 83.9%, 1H NMR (400 MHz, CD3OD) δ (ppm): 7.55 (d, J = 8.4 Hz, 2H), 7.45 (d,
J = 8.8 Hz, 2H), 4.41 (s, 1H), 3.88-3.81 (m, 3H), 3.30-3.22 (m, 3H), 2.88-2.85 (m, 2H). MS (ESI) m/z:
254 [M-H]-.
4.2.8.6. 2-(4-(Tert-butoxycarbonyl)piperazin-1-yl)-2-(4-chlorophenyl)acetic acid (13f)
White solid, yield 43.4%, 1H NMR (400 MHz, DMSO-d6) δ (ppm): 7.45 (d, J = 8.8 Hz, 2H), 7.42
(d, J = 8.8 Hz, 2H), 4.05 (s, 1H), 3.30-3.24 (m, 4H), 2.37-2.29 (m, 4H), 1.37 (s, 9H). MS (ESI) m/z: 353
[M-H]-.
4.2.8.7. Methyl 2-(4-chlorophenyl)-2-(4-methylpiperazin-1-yl)acetate (13g)
Paleyellow solid, yield 93.9%, 1H NMR (400 MHz, CD3OD-d6) δ (ppm): 7.50 (d, J = 8.4 Hz, 2H),
7.34 (d, J = 8.4 Hz, 2H), 3.88 (s, 1H), 3.31-3.30 (m, 3H), 3.14 (s, 3H), 2.74 (s, 3H), 2.65 (br, 2H). MS
(ESI) m/z: 267 [M-H]-.
4.2.8.8. 2-(4-((Tert-butoxycarbonyl)amino)piperidin-1-yl)-2-(4-chlorophenyl)acetic acid (13h)
White solid, yield 59.8%, 1H NMR (400 MHz, DMSO-d6) δ (ppm): 7.42 (s, 4H), 6.84 (d, J = 7.2
Hz, 1H), 4.04 (s, 1H), 3.26 (s, 1H), 3.05 (d, J = 11.2 Hz, 1H), 2.64 (d, J = 11.6 Hz, 1H), 2.36 (t, J = 10.8
Hz, 1H), 2.23 (t, J = 10.8 Hz, 1H), 1.75 (d, J = 6.0 Hz, 1H), 1.68 (d, J = 12.4 Hz, 1H), 1.50 (t, J = 12.0
Hz, 2H), 1.36 (s, 9H). MS (ESI) m/z: 367 [M-H]-.
4.2.9. General synthesis of compounds 14
To a solution of 13a-13h (0.4 mmol) and DIEA (2.0 mmol) in DMF (2 mL) was added EDCI (0.48
mmol) and HOBt (0.48 mmoL). The reaction mixture was stirred at 0℃ for 1 hour, followed by addition
of 4 (0.40 mmol). After completion of addition, the final mixture was stirred at room temperature for

another 12 hours, then poured into ice water (20 mL), extractd with ethyl acetate (20×2 mL), the
combined organic layer was washed with saturated aqueous sodium bicarbonate (20×3 mL) and brine
(20×2 mL), dried over sodium sulfate and solvent was evaporated under reduced pressure. The crude
product was purified by silica gel chromatography eluting with ethyl acetate /methanol (100:1~60:1) to
give 14.
4.2.9.1. 1-(4-(5-Bromo-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-2-(4-chlorophenyl)-2-
(dimethylamino)ethan-1-one (14a)
Brown solid, yield 74.4%, mp: 103~105℃, 1H NMR (400 MHz, CDCl3) δ (ppm): 12.06 (s, 1H),
8.37 (s, 1H), 7.40 (d, J = 8.5 Hz, 2H), 7.34 (d, J = 8.5 Hz, 2H), 7.29 (s, 1H), 4.26 (s, 1H), 3.95-3.91 (m,
1H), 3.88-3.62 (m, 4H), 3.59-3.50 (m, 2H), 3.33-3.19 (m, 1H), 2.32 (s, 6H), 1.26 (t, J = 7.1 Hz, 1H). 13
C
NMR (100 MHz, CDCl3) δ (ppm): 169.27, 159.67, 151.95, 150.63, 134.55, 134.21, 130.33, 129.00,
123.49, 105.36, 87.34, 71.55, 50.38, 49.59, 45.29, 43.39, 42.12. MS (ESI) m/z: 477 [M+H]+
.
4.2.9.2. 1-(4-(5-Bromo-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-2-(4-chlorophenyl)-2-
(diethylamino)ethan-1-one (14b)
Brown solid, yield 62.3%, mp: 83~87℃, 1H NMR (400 MHz, CDCl3) δ (ppm): 11.78 (s, 1H), 8.41
(s, 1H), 7.39 (d, J = 8.3 Hz, 2H), 7.34 (d, J = 8.4 Hz, 2H), 4.84 (s, 1H), 3.94-3.81 (m, 3H), 3.71-3.55 (m,
4H), 3.49-3.40 (m, 1H), 2.81 (dd, J = 13.2, 6.8 Hz, 2H), 2.63 (dd, J = 13.1, 6.7 Hz, 2H), 1.28 (s, 2H),
1.03 (t, J = 7.0 Hz, 6H). 13C NMR (100 MHz, CDCl3) δ (ppm): 170.11, 159.73, 151.91, 150.50, 135.28,
133.74, 130.66, 128.68 (2C), 123.50 (2C), 105.38, 87.34, 65.77, 50.45, 49.75, 45.30, 44.33 (2C), 41.90,
29.71, 13.04 (2C). MS (ESI) m/z: 505 [M+H]+
.
4.2.9.3. 1-(4-(5-Bromo-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-2-(4-chlorophenyl)-2-
(pyrrolidin-1-yl)ethan-1-one (14c)
Brown solid, yield 64.6%, mp: 112~115℃, 1H NMR (400 MHz, CDCl3) δ (ppm): 11.87 (s, 1H),
8.36 (s, 1H), 7.38 (dd, J = 42.1, 7.4 Hz, 4H), 4.37 (s, 1H), 3.71 (dd, J = 98.1, 45.4 Hz, 7H), 3.26 (s, 1H),
2.61 (d, J = 26.6 Hz, 4H), 1.81 (s, 4H). 13C NMR (100 MHz, DMSO) δ (ppm): 168.76, 159.55, 152.09,
151.09, 132.91, 130.84, 128.90 (2C), 124.84 (2C), 104.96, 86.24, 67.78, 51.74, 50.30 (2C), 45.24 (2C),
42.02 (2C), 23.44 (2C), 14.55. MS (ESI) m/z: 503 [M+H]+
.
4.2.9.4. 1-(4-(5-Bromo-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-2-(4-chlorophenyl)-2-(3,5-
dimethylpiperidin-1-yl)ethan-1-one (14d)
Brown solid, yield 51.0%, mp: 114~117℃, 1H NMR (400 MHz, CDCl3) δ (ppm): 11.85 (s, 1H),

8.38 (s, 1H), 7.38 (d, J = 8.5 Hz, 2H), 7.33 (d, J = 8.5 Hz, 2H), 4.39 (s, 1H), 3.95-3.71 (m, 4H), 3.70-
3.47 (m, 3H), 3.47-3.33 (m, 1H), 2.85 (t, J = 8.3 Hz, 2H), 1.83-1.62 (m, 4H), 1.54-1.43 (m, 1H), 1.25 (s,
2H), 0.83-0.78 (m, 6H), 0.55-0.47 (m, 1H). 13C NMR (100 MHz, CDCl3) δ (ppm): 169.38, 159.70, 151.92,
150.65, 133.82, 130.54, 128.72, 123.23, 105.30, 87.52, 71.53, 59.43, 58.51, 50.39, 49.70, 45.26, 42.10,
31.94, 31.34, 31.15, 29.71, 29.38, 22.71, 19.61, 14.15. MS (ESI) m/z: 545 [M+H]+
.
4.2.9.5. 1-(4-(5-Bromo-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-2-(4-chlorophenyl)-2-
morpholinoethan-1-one (14e)
Brown solid, yield 70.0%, mp: 110~114℃, 1H NMR (400 MHz, CDCl3) δ (ppm): 11.91 (s, 1H),
8.38 (s, 1H), 7.40 (d, J = 8.5 Hz, 2H), 7.35 (d, J = 8.5 Hz, 2H), 7.28 (s, 1H), 4.00-3.62 (m, 10H), 3.56
(dd, J = 16.1, 11.5 Hz, 2H), 3.30 (dd, J = 10.0, 7.3 Hz, 1H), 2.55 (s, 4H). 13C NMR (100 MHz, CDCl3)
δ (ppm): 168.65, 159.67, 151.92, 150.64, 134.38, 133.47, 130.45, 129.06, 123.33, 105.36, 87.45, 71.15,
66.89, 60.43, 51.75, 50.38, 49.58, 45.30, 42.15. MS (ESI) m/z: 519 [M+H]+
.
4.2.9.6. Tert-butyl 4-(2-(4-(5-bromo-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-1-(4-
chlorophenyl)-2-oxoethyl)piperazine-1-carboxylate (14f)
Brown solid, yield 76.2%, mp: 131~135℃, 1H NMR (400 MHz, CDCl3) δ (ppm): 12.02 (s, 1H),
8.30 (s, 1H), 7.40-7.23 (m, 4H), 7.20 (s, 1H), 4.30 (s, 1H), 3.93-3.14 (m, 12H), 2.58-2.28 (m, 4H), 1.36
(s, 10H). MS (ESI) m/z: 618 [M+H]+
.
4.2.9.7. 1-(4-(5-Bromo-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-2-(4-chlorophenyl)-2-(4-
methylpiperazin-1-yl)ethan-1-one (14g)
Brown solid, yield 51.6%, mp: 85~87℃, 1H NMR (400 MHz, CDCl3) δ (ppm): 8.37 (s, 1H), 7.39
(d, J = 8.5 Hz, 2H), 7.33 (d, J = 8.5 Hz, 2H), 7.26 (s, 1H), 4.31 (s, 1H), 3.95-3.46 (m, 8H), 3.32 (dd, J =
10.0, 7.2 Hz, 1H), 2.54 (d, J = 26.6 Hz, 7H), 2.28 (s, 3H). 13C NMR (100 MHz, CDCl3) δ (ppm): 168.87,
159.67, 151.96, 150.68, 134.12, 133.92, 130.37, 128.91, 123.42, 105.37, 87.31, 71.28, 54.95, 51.17,
50.38, 49.63, 46.08, 45.87, 45.26, 42.19, 10.76. MS (ESI) m/z: 532 [M+H]+
.
4.2.9.8. Tert-butyl (1-(2-(4-(5-bromo-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-1-(4-
chlorophenyl)-2-oxoethyl)piperidin-4-yl)carbamate (14h)
Brown solid, yield 54.5%, mp: 140~143℃, 1H NMR (400 MHz, CDCl3) δ (ppm): 11.63 (s, 1H),
8.36 (s, 1H), 7.37 (d, J = 8.5 Hz, 2H), 7.33 (d, J = 8.5 Hz, 2H), 4.42 (d, J = 52.5 Hz, 2H), 3.93-3.40 (m,
9H), 3.32 (s, 1H), 3.00-2.75 (m, 2H), 2.32 (s, 1H), 2.19-1.99 (m, 2H), 1.91 (t, J = 14.5 Hz, 2H), 1.58-
1.35 (m, 12H). MS (ESI) m/z: 632 [M+H]+
.

4.2.10. General synthesis of compounds 15a-15b
Compounds 14a-14b (0.1 mmol) was suspended in methanol (2 mL), after addition of the 4M HCl
in dioxane (6 ml), the reaction mixture was stirred for 48 hours at room temperature. The precipitate was
filtrated, the obtained solid was recrystallized from methanol to give the desired 15a-15b.
4.2.10.1. 1-(4-(5-Bromo-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-2-(4-chlorophenyl)-2-
(piperazin-1-yl)ethan-1-one hydrochloride (15a)
White solid, yield 98.9%, mp: 150~155℃, 1H NMR (400 MHz, D2O) δ (ppm): 8.10 (s, 1H), 7.43
(s, 5H), 7.35 (s, 1H), 5.54 (s, 1H), 4.05-3.94 (m, 1H), 3.83 (qd, J = 12.9, 4.5 Hz, 2H), 3.76-3.68 (m, 1H),
3.67-3.30 (m, 15H), 3.29-3.16 (m, 3H), 2.93-2.82 (m, 1H). 13C NMR (100 MHz, D2O) δ (ppm): 164.96,
154.30, 146.52, 143.05, 137.33, 131.14, 130.53, 126.65, 125.98, 103.02, 88.91, 69.29, 49.61, 48.51,
48.08, 44.45, 42.06, 40.80. MS (ESI) m/z: 518 [M+H]+
.
4.2.10.2. 2-(4-Aminopiperidin-1-yl)-1-(4-(5-bromo-7H-pyrrolo[2,3-d]pyrimidin-4-yl)piperazin-1-yl)-2-
(4-chlorophenyl)ethan-1-one hydrochloride (15b)
White solid, yield 99.2%, mp: 147~151℃, 1H NMR (400 MHz, D2O) δ (ppm): 8.11 (s, 1H), 7.45
(s, 5H), 7.37 (s, 1H), 5.52 (s, 1H), 4.09-3.96 (m, 1H), 3.97-3.28 (m, 14H), 3.26-2.81 (m, 6H), 2.27-2.21
(m, 3H), 2.05-1.76 (m, 3H). 13C NMR (100 MHz, D2O) δ (ppm): 164.76, 154.54, 151.00, 146.71, 143.36,
137.35, 130.49, 130.42, 126.57, 125.95, 103.14, 88.81, 66.52, 62.47, 49.57, 49.02, 48.49, 45.23, 44.46,
44.28, 42.10. MS (ESI) m/z: 532 [M+H]+
.
4.3. In vitro Akt1 kinase activity assay
The in vitro Akt1 kinase activity was evaluated via an HTRF assay (LANCER○) using the Akt kinase
kit (Cisbio Bioassays, No.62ST3PEB) in 384-well plates. Each well was added Akt1, STK
Substrate-biotin, tested compounds and ATP in kinase buffer (50 mM HEPES, 5 mM MgCl2, 1 mM
DTT, 0.02% NaN3 and 0.01% BSA, pH = 7.0) in sequence, then incubated for 45 min at 25°C.
Finally, Sa-XL665 and STK Ab-Cryptate were added to stop the enzymatic step and incubated for 2
hours to finish the detection process. The ratio (665 nm/620 nm) was obtained using a microplate
reader (Perkin Elmer, USA).
4.4 Z'-LYTE Kinase Assay
The reaction was conducted in a 384-well plate with 10 μL reaction volume per well containing 2
μM Ser/Thr 06 peptide substrate in 50 mM HEPES pH 7.5, 0.01% BRIJ-35, 10 mM MgCl2, 1 mM EGTA
and appropriate amount of AKT kinases with a serial 3-fold dilution of tested compound. The final

reaction concentration of ATP was 75 μM for Akt1, 200 μM for Akt2 and 100 μM for Akt3. After 1 hour
kinase reaction incubation, 5 µL of a 1:4096 dilution of development reagent A is added, reaction was
developed for another 1 hour and then terminated. Fluorescein FRET signal were monitored at 445 and
520 nm, respectively. The fluorescence ratio was calculated according to the manufacturer’s protocol
(ThermoFisher, USA).
4.5. Cell lines and primary MCL cells
MCL cell lines Granta519, Maver-1, Rec-1, Z-138, Mino and Jeko-1 were purchased from the
American Type Culture Collection (ATCC). Human MCL cells were purified from the apheresis of MCL
patients after obtaining informed consent and approval by the Institutional Review Board at The
University of Texas MD Anderson Cancer Center. Patient peripheral blood mononuclear cells (PBMCs)
were isolated by Ficoll-Hypaque density centrifugation and stained for human anti-CD5 and human anti-
CD20 antibodies, CD5 and CD20 double positive cells representing MCL cell population were detected
by Novocyte Flow Cytometer (ACEA Bioscience, USA). Isolated samples with more than 95%
CD5+CD20+ MCL cells were used without further purification. Cells were further cultured in RPMI-
1640, supplemented with 10% heat-inactivated fetal bovine serum, 2% HEPES buffer and penicillin
(10,000 units/mL; Sigma), streptomycin (10 mg/mL; Sigma).
4.6. Cell proliferation assay
Cell proliferation assay was performed on a panel of MCL cell lines and primary MCL cells using
the CellTiter-Glo Luminescent cell viability assay kit (Promega) following the manufacturer’s protocol.
In short, cells were plated in 96-well plates at a density of 1×104 cells/well for cell lines and 12.5×104
cells/well for primary MCL cells, then treated with DMSO and different concentrations of the
synthesized compounds and incubated in a humidified atmosphere with 5% CO2 at 37°C for 72 hours on
MCL cell lines and 24 hours on primary MCL cells. IC50 values were calculated using the Graphpad
prism 6 software.
4.7. Cell apoptosis assay
Apoptosis was quantified by Annexin V/Propidium Iodide (PI)-binding assay. Cells were seeded in
6-well plates with 1 μM, 2.5 μM and 5 μM of 8 and 14g for 24 hours. Treated cells were washed twice
with cold phosphate-buffered saline (PBS) and then resuspended in100 µL binding buffer, to which 2
µL of Annexin V-FITC and 5 µL of PI were added. The samples were gently vortexed and incubated for
15 minutes at room temperature in the dark. After addition of 200 μL binding buffer, samples were

immediately analyzed by flow cytometry using a NovoCyte Flow Cytometer (ACEA Biosciences, San
Diego, CA). The number of apoptotic cells was determined using the NovoExpress software.
4.8. Cell cycle assay
Cell cycle arrest was measured using PI staining by flow cytometry. Cells was seeded in 6-well
plates with 1 μM and 2.5 μM of 8 and 14g for 24 hours, then cells were harvested, washed twice with
cold PBS, and subsequent fixing in cold 70% ethanol overnight at 4 ℃. Then samples were washed twice
with PBS, followed by further treatment with 50 uL of 100 ug/mL ribonuclease and 200 uL 50 ug/mL
PI, and finally analyzed by NovoCyte Flow Cytometer (ACEA Biosciences, San Diego, CA).
4.9. Western blotting
Jeko-1 cells and patient primary cells were cultured with different concentrations of 8 and 14g for
24 hours. Then cells were harvested and lysed in a lysis buffer (Cell Signaling, Danvers, MA). The cell
lysates were kept on ice for 30 minutes and centrifuged at 12,000×rpm for 20 minutes at 4oC. The protein
concentration was determined by the Bradford assay (Bio-Rad, Hercules, CA). Twenty micrograms of
sample proteins were mixed with the loading buffer and separated by 10% SDS-PAGE. The proteins
were then transferred onto methanol equilibrated PVDF membrane (BIO-RAD Laboratories, 162-0177),
which was blocked for 1 hour in 5% nonfat dry milk in TBST (BD Bioscience, San Jose, CA). The
membranes were incubated with a primary antibody overnight at 4 ℃. Secondary antibodies were added
for 1 hour at room temperature. Finally, the membrane was visualized by ECL (Perkin Elmer Life
Sciences, NE104001EA). Antibodies against PARP, cleaved caspase 3, p-Akt-308, p-Akt-473, pan-Akt,
p-GSK3β, total- GSK3β and GAPDH were obtained from Cell Signaling.
4.10. Molecular docking
Molecular docking was performed using the Sybyl 2.0 software package and the Akt1 crystal
structure (PDB: 3MV5) was retrieved from the Protein Data Bank. Protein preparation was performed
by extracting the ligand, removing water molecules, adding hydrogen atoms and assigning AMBER7
FF99 charges to the protein. Compounds 8 and 14g was docked into Akt1 and the hydrogen bonds and
hydrophobic interactions were observed in the model, the best conformation with the highest CScore was
selected for interaction analysis.
Acknowledgement
This work was supported by the Key Research and Development Project of Shandong Province
(Nos. 2017CXGC1401 and 2019GSF108038)
Conflict of interest

The authors declare no conflict of interest.
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Declaration of interests
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.
☐The authors declare the following financial interests/personal relationships which may be
considered as potential competing interests:

Highlights



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Further exploration of pyrrolopyrimidine analogues as Akt inhibitors
High enzymatic potency and antiproliferative effects in lymphoma cells
Induction of cell apoptosis and G2/M cell cycle arrest
Suppressed phosphorylation level of Akt downstream targets GSK3β and S6