Discovery of 2-hydroxyarbekacin, a new aminoglycoside antibiotic with reduced nephrotoxicity

Full text of DOI:10.1038/ja.2017.60

The Journal of Antibiotics (2017), 1–3
2017 Japan Antibiotics Research Association All rights reserved 0021-8820/17
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COMMUNICATION TO THE EDITOR
Discovery of 2-hydroxyarbekacin, a new aminoglycoside
antibiotic with reduced nephrotoxicity
The Journal of Antibiotics advance online publication, 28 June 2017; doi:10.1038/ja.2017.60
The emergence and spread of bacteria with 2-hydroxyarbekacin (2-OH-ABK) was also of the 2-hydroxyl group to yield 12 (89%).
resistance to antibacterial drugs in recent of interest from the viewpoint of its potential Acidic hydrolysis quantitatively gave the
years is now considered a significant threat antibacterial activity.
deacetonated derivative 13, which was treated
to global public health and the world
As shown in Scheme 1, the synthesis of with acetic acid–acetic anhydride–sulfuric
economy.1,2 In particular, the severe bacterial 2-OH-ABK was performed by the condensa- acid (50:50:1), thereby affording the glycosyl
infections caused by methicillin-resistant tion of 1-N-[(S)-4-benzyloxycarbonylami- acetate 14 (92%). Bromination of 14 with
Staphylococcus aureus (MRSA) and Pseudomo- no-2-benzyloxybutylyl]-2-benzyloxy-3,2′,6′-
nas aeruginosa have become serious clinical tris(N-benzyloxycarbonyl)-3′,4′-dideoxynea-
TiBr4 in CH2Cl2–EtOAc (9:1) gave the corre-
sponding α-bromide 15 (97%) as a syrup, 1H
problems because of increased antimicrobial mine (9), which contains an equatorial NMR (400 MHz, CDCl3) δ 3.43 (dd, 1H,
resistance, as observed for vancomycin- hydroxyl group at C-2 and phenyl 4,6-di-O- H-2), 3.98 (t, 1H, H-3), 4.92 (t, 1H, H-4) and
resistant S. aureus and multi-drug resistant acetyl-3-azido-2-O-benzyl-3-deoxy-1-thio-α-
6.32
(d,
1H,
H-1);
J1,2 = 3.8 Hz
P. aeruginosa.3 Arbekacin4 (ABK), an D-glucopyranoside (16).
and J2,3 = J3,4 = 10 Hz. Treatment of 15 with
trimethyl(phenylthio)silane in the presence of
trimethylsilyl trifluoromethanesulfonate in
CH2Cl2, gave 1-α-thiophenyl glycoside 16
(76%) as crystals, 1H NMR (400 MHz,
CDCl3) δ 1.99 and 2.13 (each s, 3H, Ac),
3.79 (dd, 1H, H-2), 3.86 (t, 1H, H-3), 3.96
(t, 1H, H-6a), 4.22 (dd, 1H, H-6b), 4.46
(ddd, 1H, H-5), 4.72 and 4.75 (ABq, 2H,
Jgem = 12 Hz, CH2Ph), 4.83 (t, 1H, H-4) and
5.59 (d, 1H, H-1); J1,2 = 5 Hz, J2,3 = J3,4
J4,5 = 10 Hz, J5,6a = 2.5 Hz, J5,6b = 5.5 Hz and
J6a,6b = 12.5 Hz.
Condensation of 9 and 16 was carried
out successfully in CH2Cl2 in the presence
of N-iodosuccinimide, trifluoromethanesulfo-
nic acid and 4A molecular sieves to give 17
Acidic hydrolysis of the natural product,
2-hydroxygentamicin C1a10, gave the pseudo-
disaccharide, 3′, 4′-dideoxy-2-hydroxynea-
mine (1) (85%, as the bis-carbonate).
N-Benzyloxycarbonylation of 1 gave 2 in
quantitative yield, which was then treated
with 1,1-dimethoxycyclohexane in the
presence of pyridinium p-toluenesulfonate
to yield the cyclohexylidene derivative 3
(85%). Next, 2-O-benzylation gave 4 (83%),
followed by removal of its acetal protecting
group in an aqueous acidic solution to
provide the 5, 6-diol 5 (93%). Treatment of
aminoglycoside antibiotic, is efficacious
against MRSA, and is commonly used to
treat infected patients in the clinical setting.
This semi-synthetic antibiotic is stable
toward AAC(6′)-APH(2″),
a bifunctional
enzyme present in MRSA, and is also effective
against almost all antibiotic-resistant bacteria
that produce aminoglycoside-modifying
enzymes.5 However, despite its superior anti-
bacterial activity and stability toward
aminoglycoside-modifying enzymes, treat-
ment with ABK is limited because of its
adverse effect on the kidneys.6,7 Therefore,
=
we have conducted synthetic studies on a 5 with NaH (6 equiv. relative to 5) in DMF at
number of ABK derivatives in an effort to room temperature gave the N,O-carbonyl
decrease the associated toxicity.8,9
derivative 6 (95%), which was retreated with
It is generally known that the nephrotoxi- NaH (1 equiv. relative to 6) in aqueous (69% based on 9). After removal of the acetyl
city of streptomycin is relatively low among dioxane at 80 °C to yield the 1-amino group, the resulting product 18 (87%) was
aminoglycoside antibiotics. Streptomycin derivative 7 (77%). Subsequent introduction treated with hydrogen in the presence of
retains N-amidinated streptamine (strepti- of an amino acid side-chain to 7, which palladium black to reduce the azido group
dine) as a constituent aminocyclitol, and possesses a free amino group at C-1, was and remove the benzyl and benzyloxycarbonyl
kanamycins such as ABK contain 2-deoxy- successfully accomplished by utilizing the groups to yield, after purification by Amber-
streptamine as
a constituent component active ester 8 according to conventional lite CG-50 resin column chromatography, a
(Figure 1). By comparing the structure of methodology11 to afford the 1-N-acyl deriva- solid of 2-OH-ABK (72%) as the carbonate,
these two aminocyclitols, we focused on an tive 9 (76%), H NMR (400 MHz, pyridine- [α]D25+96 (c 1, H2O); HR-MS: m/z calcula-
1
equatorial hydroxyl group at the C-2 position d5) δ 2.28 (m, 2H, H-3″′) and 5.65 (d, 1H, tion for C22H45N6O11 (M+H)+ 569.3141,
1
and were interested in the relationship
between the presence of this hydroxyl
found 569.3151; H NMR (500 MHz, DCl −
D2O, pD 2) δ 1.56 (m, 1H, H-4′ax), 2.14
J = 2.9 Hz, H-1′).
Next, the glycosyl donor 16 was prepared.
group and nephrotoxicity. Also, because the Acetalization of methyl 3-azido-3-deoxy-α, (ddt, 1H, H-3″′b), 3.06 (dd, 1H, H-6′a), 3.12
modification at the C-2 position of the ABK β-D-glucopyranoside gave the (t, 2H, H-4″′), 3.21 (dd, 1H, H-6′b), 3.35
(10)12
derivative is not known, the synthesis of acetonide 11 (96%), followed by benzylation (apparent t, 1H, H-3″), 3.37 (t, 1H, H-3), 3.52

Communication to the Editor
2
NH
NHCNH2
NH
1
OH
HO
3
2
OH
OH
H2NCHN
4
O
HO
H2N
H2N
6'
O
O
1"
1'
NH2
O
OH
HO
CHO
O
O
HN
2
HO
Me
NH2
3
1
HO
O
HO
HO
O
H2N
O
NHMe
HO
Streptomycin
Arbekacin (ABK)
Figure 1 Structures of streptomycin and arbekacin, and the points of interest. A full colour version of this figure is available at the Journal of Antibiotics
journal online.
Scheme 1 Synthesis of 2-hydroxyarbekacin (2-OH-ABK).
(m, 1H, H-2′), 3.63 (t, 1H, H-4″), 3.72
(dd, 1H, H-2″), 3.76 (t, 1H, H-2), 4.15 activity in vitro against both S. aureus and P. (6′)-I] they were 4 and 16, respectively.
(m, 1H, H-5′), 4.28 (dd, 1H, H-2″′), 5.09 aeruginosa. The MICs (μg ml− l
of
The nephrotoxicity of 2-OH-ABK was
2-OH-ABK exhibited excellent antibacterial and against P. aeruginosa PAO1/315 [AAC
)
(d, 1H, H-1″) and 5.75 (d, 1H, H-1′); 2-OH-ABK and ABK against S. aureus examined using an N-acetyl-β-D-glucosamini-
J1,2 = J2,3 = J3,4 = 10.5 Hz, J1′,2′ = 3.5 Hz, J5′,6′ RN4220 (MSSA) were 0.5 and 0.5, against S. dase (NAG) assay13 in vitro and a rat in vivo
a = 7.5 Hz, J5′,6′b = 3.5 Hz, J6′a,6′b = 13.5 Hz, aureus RN4220/pCR1948 [AAC(6′) − APH test. As shown in Figure 2a, the amount of
J1″,2″ = 3.8 Hz, J2″,3″ = 11 Hz, J3″,4″ = 10 Hz, (2″)] they were 0.5 and 2, and against clinical NAG released from LLC-PK1 cells in the
J2″′, 3″′a = 9.5 Hz, J2″′, 3″′b = 4 Hz and J3″′a, 4″′
=
isolates of MRSA (n = 77) they were 0.25–2 presence of 2-OH-ABK was lower than that
J3″′b, 4″′ = 7.5 Hz. Anal calcd for C22H44N6O11 and 0.25–4, respectively. Furthermore, the of ABK at any dose (2.5, 5 and 10 μM),
2H2CO3 H2O: C 40.56, H 7.09, N 11.83. MICs (μg ml− 1) of 2-OH-ABK and ABK indicating a tendency of reduction in nephro-
Found: C 40.44, H 7.10, N 11.53.
against P. aeruginosa PAO1 were 2 and 4, toxicity for 2-OH-ABK. Furthermore,
The Journal of Antibiotics

Communication to the Editor
3
Yoshiaki Takahashi1, Eijiro Umemura1,
Yoshihiko Kobayashi1, Shoichi Murakami2,
Toru Nawa2, Akihiro Morinaka2,
3.00
2.50
2.00
1.50
1.00
0.50
0.00
Toshiaki Miyake1 and Masakatsu Shibasaki1
1Institute of Microbial Chemistry
P<0.01
2
(BIKAKEN), Tokyo, Japan and Meiji Seika
P<0.05
Pharma Co., Ltd., Yokohama, Japan
E-mail: takashow@bikaken.or.jp
control
2.5
5
10
2.5
5
10
1
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Figure 2 Nephrotoxicity of 2-hydroxyarbekacin (2-OH-ABK) in vitro and in vivo. (a) N-acetyl-β-D-
glucosaminidase (NAG) assay for 2-OH-ABK. NAG release from LLC-PK1 cells, grown on porous culture
inserts as individual confluent monolayers, was determined in the in vitro nephrotoxic screening. NAG
activity in the serum-free apical conditioned medium was quantified using sodio-m-
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Figure 2b shows the measurements of urea nephrotoxicity was lower than that of ABK.
9
nitrogen and creatinine in the blood from the
drug-treated rats. In this test, although
significant increases in blood urea nitrogen
and creatinine values were observed in rats
treated with ABK, no such increases in these
values were observed for the 2-OH-ABK-
treated rats. Additionally, the histopathologi-
cal renal tubular epithelial necrosis observed
with ABK was not observed in the 2-OH-
ABK-treated rats.
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CONFLICT OF INTEREST
The authors declare no conflict of interest.
In summary, a novel aminoglycoside anti-
biotic, 2-OH-ABK, was created and charac-
terized in our drug discovery program on
ACKNOWLEDGEMENTS
We are grateful to Dr Minoru Yonezawa at the
Meiji Seika Pharma Co., Ltd. for providing valuable
next-generation antibiotics. Notably, its information on the biological studies.
The Journal of Antibiotics