Design, synthesis and structure-activity relationships of novel macrolones: Hybrids of 2-fluoro 9-oxime ketolides and carbamoyl quinolones with highly improved activity against resistant pathogens

Article abstract of DOI:10.1016/j.ejmech.2019.02.073

Constitutively erythromycin-resistant apathogens are more difficult to address than inducibly resistant and efflux-resistant strains. Three series of the 4th generation 2-fluoro 9-oxime erythromycin ketolides were synthesized and evaluated. Incorporation of substituted heteroaryl groups (a - m), in contrast to previously reported the unsubstituted heteroaryl groups, proved to the beneficial for enhancement of the activities of the 9-propgargyl ketolide 8 series and the 9-allyl ketolide 14 series. But these aryl groups (a - m) cannot supply the resulting compounds 8 and 14, unlike corresponding the 6-allyl ketolide 20 series, with activity against constitutively resistant Streptococcus pneumoniae. However, hybrids of macrolides and quinolones (8, 14 and 20, Ar = n - t) exhibited not only high activities against susceptible, inducibly erm-mediated resistant, and efflux-mediated resistant strains, but also significantly improved potencies against constitutively resistant Streptococcus pneumoniae and Streptococcus pyogenes. The capacity was highlighted by introduction of newly designed carbamoyl quinolones (q, r, s and t) rather than commonly seen carboxy quinolones (o and p) as the pharmacophores. Structure-activity relationships and molecular modelling indicated that 8r, 14r and 20q may have different binding sites compared to current erythromycins. Moreover, 8r, 14r and 20q have 2.5–3.6 times prolonged half-life and 2.3- to 2.6-fold longer mean residence time in vivo over telithromycin. These findings pave the way for rational design of novel non-telithromycin macrolides that target new binding sites within bacterial ribosomes.

Full text of DOI:10.1016/j.ejmech.2019.02.073

Accepted Manuscript
Design, synthesis and structure-activity relationships of novel macrolones: Hybrids
of 2-fluoro 9-oxime ketolides and carbamoyl quinolones with highly improved activity
against resistant pathogens
Cong-Xuan Ma, Wei Lv, Ya-Xin Li, Bing-Zhi Fan, Xu Han, Fan-Sheng Kong, Jing-
Chao Tian, Mark Cushman, Jian-Hua Liang
PII:
S0223-5234(19)30205-3
DOI:
https://doi.org/10.1016/j.ejmech.2019.02.073
EJMECH 11165
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 30 January 2019
Revised Date: 27 February 2019
Accepted Date: 27 February 2019
Please cite this article as: C.-X. Ma, W. Lv, Y.-X. Li, B.-Z. Fan, X. Han, F.-S. Kong, J.-C. Tian, M.
Cushman, J.-H. Liang, Design, synthesis and structure-activity relationships of novel macrolones:
Hybrids of 2-fluoro 9-oxime ketolides and carbamoyl quinolones with highly improved activity against
resistant pathogens, European Journal of Medicinal Chemistry (2019), doi: https://doi.org/10.1016/
j.ejmech.2019.02.073.
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ACCEPTED MANUSCRIPT
Graphical abstract
Design, Synthesis and Structure-activity Relationships of Novel
Macrolones: Hybrids of 2-Fluoro 9-Oxime Ketolides and Carbamoyl
Quinolones with Highly Improved Activity against Resistant Pathogens
a
b
c
a
c
d
Cong-Xuan Ma, Wei Lv, Ya-Xin Li, Bing-Zhi Fan, Xu Han, Fan-Sheng Kong,
c
b
a, c *
Jing-Chao Tian, Mark Cushman, Jian-Hua Liang

ACCEPTED MANUSCRIPT
Design, Synthesis and Structure-activity Relationships of Novel
Macrolones: Hybrids of 2-Fluoro 9-Oxime Ketolides and Carbamoyl
Quinolones with Highly Improved Activity against Resistant Pathogens
a
b
c
a
c
d
Cong-Xuan Ma, Wei Lv, Ya-Xin Li, Bing-Zhi Fan, Xu Han, Fan-Sheng Kong,
c
b
a, c *
Jing-Chao Tian, Mark Cushman, Jian-Hua Liang
a
School of Chemistry and Chemical Engineering, Beijing Institute of Technology, Beijing 100081, China
b
Department of Medicinal Chemistry and Molecular Pharmacology, College of Pharmacy, and the Purdue Center
for Cancer Research, Purdue University 47907 USA
c
School of Life Science, Beijing Institute of Technology, Beijing 100081, China
d
Beijing Increasepharm Safety & Efficacy Co. Ltd, Beijing 102206, China
_
*
_________________
To whom correspondence should be addressed. J.-H. Liang: E-mail: ljhbit@bit.edu.cn
1

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Abstract: Constitutively erythromycin-resistant pathogens are more difficult to address than
th
inducibly resistant and efflux-resistant strains. Three series of the 4 generation 2-fluoro 9-oxime
erythromycin ketolides were synthesized and evaluated. Incorporation of substituted heteroaryl
groups (a - m), in contrast to previously reported the unsubstituted heteroaryl groups, proved to
the beneficial for enhancement of the activities of the 9-propgargyl ketolide 8 series and the 9-allyl
ketolide 14 series. But these aryl groups (a - m) cannot supply the resulting compounds 8 and 14,
unlike corresponding the 6-allyl ketolide 20 series, with activity against constitutively resistant
Streptococcus pneumoniae. However, hybrids of macrolides and quinolones (8, 14 and 20, Ar = n
-
t) exhibited not only high activities against susceptible, inducibly erm-mediated resistant, and
efflux-mediated resistant strains, but also significantly improved potencies against constitutively
resistant Streptococcus pneumoniae and Streptococcus pyogenes. The capacity was highlighted by
introduction of newly designed carbamoyl quinolones (q, r, s and t) rather than commonly seen
carboxy quinolones (o and p) as the pharmacophores. Structure-activity relationships and
molecular modelling indicated that 8r, 14r and 20q may have different binding sites compared to
current erythromycins. Moreover, 8r, 14r and 20q have 2.5 - 3.6 times prolonged half-life and
2
.3- to 2.6-fold longer mean residence time in vivo over telithromycin. These findings pave the
way for rational design of novel non-telithromycin macrolides that target new binding sites within
bacterial ribosomes.
Key Words: erythromycin; ciprofloxacin; multi-drug resistance; ribosome; erythromycin
ribosomal methylation; macrolide efflux
2

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1
. Introduction
Erythromycin (generally referred to as erythromycin A, Fig. 1) has been clinically used for
safely curing upper and lower respiratory tract infection, especially community-acquired bacterial
pneumonia, for more than 60 years. Erythromycin is long viewed as being a bacteriostatic agent
due to its global arrest of nascent-peptide release from bacterial ribosomes. [1] The hydrogen
6
bonding formed between the 5-O-desosamine of erythromycin and the N of the ribosomal RNA
base A2058 within the nascent peptide exit tunnel (NPET) affords the primary affinity to keep
erythromycin binding firmly to the narrowest site adjacent to the peptidyl transferase center (PTC).
[2] In the 1980s, clarithromycin and azithromycin (Fig. 1), the second-generation erythromycins,
were designed to address the intrinsic acid-instability of erythromycin. Since the late of the 1990s,
antibacterial resistance has arisen worldwide, and particularly prevalent macrolide-resistant
Streptococcus pneumoniae and methicillin-resistant Staphylococcus aureus (MRSA) in Asian
countries has caused a serious threat to public health. [3, 4] The resistant bacteria significantly
diminish the affinity of erythromycin, clarithromycin and azithromycin to bacterial ribosomes by
acquiring an erm (erythromycin ribosomal methylation) gene, which results in mono- or
6
dimethylation on the N of A2058 in the presence of erythromycins (called inducible phenotype)
or in the absence of erythromycins (denoted constitutive phenotype).
<
Figure 1>
Agouridas and Denis synthesized the 3-keto erythromycin derivatives, named ketolides. [5]
The structure-activity relationships indicated that one sugar moiety at the 3-O position, namely
3
-O-cladinose, is actually responsible for the induction of the erm-mediated resistance and mef
(macrolide efflux)-mediated resistance, and introduction of 3-keto prevents both of these types of
the resistance. It was noted that the ketolides that possess aromatic heterocycles linked to the
aglycone via an appropriate spacer are endowed with extra potencies against constitutively
resistant S. pneumoniae. [5, 6] Later, the high-resolution X-ray crystal structures of telithromycin
in complex with bacterial ribosomes clarified that the aryl group at the end of the sidechain at the
11-postion supplies telithromycin with the capacity to π-π interact at a secondary binding site, i.e.
A752. [7, 8] Telithromycin (Fig. 1) is the first marketed third-generation ketolide, and its success
has triggered intensive investigations on telithromycin-like ketolides (Fig. 2). [4, 9-11]
3

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<
Figure 2>
Unfortunately, it was reported that telithromycin is related to several severe adverse events
including irreversible liver damage. For this reason, usage of telithromycin for simple infections
was withdrawn by the FDA, but an indication for curing serious community-acquired bacterial
pneumonia survived with a black box warning. It was suspected that either the newly introduced
11-sidechain or the 3-keto could be the cause. [12, 13] The uncertainty and potential high risk
forced Abbott, Merck, Johnson & Johnson, Pfizer, Kosan and other pharmaceutical companies to
terminate or halt their ketolide research programs. [14] As a result, telithromycin remains as a
unique marketed macrolide with activity against macrolide-resistant bacteria. Recent research has
suggested that telithromycin-associated adverse effects may be attributed to the structural
similarity between nicotine (i.e. 3-(1-methyl-2-pyrrolidyl)pyridine) and the terminal aryl group
(pyridyl imidazole), which leads to an unexpected off-target interaction with nicotinic
acetylcholine receptors. [15] On the other hand, recent advances in macrolide synthesis and the
present therapeutic crisis posed by antibiotic-resistant bacteria have kindled renewed interest
macrolide development. [14, 16, 17]
In addition to telithromycin-like derivatives, cethromycin and modithromycin (Fig. 3),
featuring a sidechain attached to the 6-, or 6,11-positions, suggested an alternative structural
modification of erythromycins for probing non-A752 binding sites. [18-20] The 9-oxime ketolides
are an important subfamily. [21] We previously reported some potent 9-oxime ether ketolides (1
and 2, Fig. 4) that were discovered by structure-activity relationship (SAR)-aided drug design. [22,
2
3] These compounds, featuring a heteroaryl moiety attached to the 9-oxime, showed significantly
improved antibacterial activity over clarithromycin against some Gram-positive bacteria such as
inducibly erm-encoded resistant or mef-mediated resistant Staphylococcus aureus, Streptococcus
pneumoniae, Streptococcus pyogenes and Staphylococcus epidermidis. Molecular modelling
suggested that the ketolides 1 and 2 may interact with the rRNA base A2062 within the NPET. [23]
The NPET was long viewed as an inert passage tunnel with a 10 nm length and 1 - 2 nm width,
but recent research strongly suggested it could be a hub for nascent protein-based translation
regulation. [24] Importantly, the A2062 acts as a gate to selectively release the nascent peptides
because the A2062 is flexible and therefore some nascent peptides with specific amino residues
4

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can make it reorient to open up the tunnel space to allow for passage. [25] The selective release of
specific proteins was cited as the cause of the observed bactericidal activity of telithromycin. [1,
2
5] In contrast, quinolones exampled by ciprofloxacin target topoisomerase IV and DNA gyrase
and exert bactericidal activity by inhibiting DNA replication and transcription. Nowadays,
-carboxyquinolone is extensively utilized as the core for design of new bactericidal chemotypes.
26, 27]
3
[
<
<
Figure 3>
Figure 4>
Despite the great amount of effort already expended on macrolides, there are reasons that
support continuous studies on 2-fluoro versions of non-telithromycin analogues, namely 9-oxime
th
ether ketolides, including: (i) further evaluation of the so-called the 4 generation 2-fluoro
ketolides since the 2-fluorine substituent inhibits the tautomerization involving 2-H and 3-keto and
rd
affords a pure ketolide in contrast to the 3 generation erythromycins exampled by telithromycin;
[15, 28, 29] (ii) inability of current 9-oxime ether ketolides to address constitutively high-level
resistant S. pneumoniae, to which additional high affinity, especially via hydrogen bonding, are
required for new erythromycins in order to compensate for the loss of the primary interaction; (iii)
inefficiencies of current 9-oxime ether ketolides against Gram-negative bacteria such as
Haemophilus influenzae and Moraxella catarrhalis and the resulting narrow antibacterial
spectrum; (iv) finding metabolically stable and highly potent 2-fluoro 9-oxime ketolides that may
provide structural insight into new non-telithromycin modes of action that can suggest a non-A752
binding site within the NPET. For these reasons, novel 2-fluoro 9-oxime ether ketolides are highly
desired to broaden comprehensive understanding of the SAR of macrolides that target rRNA bases
within the NPET.
2
. Results and Discussion
2
.1 Molecular design
Non-telithromycin macrolides and their potential binding sites have provided a focal point for
much of our recent research. [22, 23, 30-33] Previous SAR studies focused on unsubstituted
monocyclic-, bicyclic- or bi-aryl groups (thienyl, pyridyl, pyrimidyl, indolyl, quinolyl, isoquinolyl,
5

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pyridyl-thienyl, imidazolyl-phenyl and benzoyl-thienyl) anchored at the end of the 9-oxime ether
ketolides, which are exampled by compounds 1 and 2 in Fig. 4. [22, 23] For the present study, we
hypothesized that acquisition of additional hydrogen bonding and strengthened π-π stacking forces
would restore the antibacterial activity of macrolides against constitutively resistant
microorganisms, and thus installing a variety of substituents (for example, electro-donating,
electron-withdrawing, hydrophobic and hydrophilic groups) onto aryl groups (a - m, Chart 1) may
increase the affinities of macrolides with potential binding sites within the NPET. Next, as
fastidious Gram-negative strains are protected by outer biofilms composed of lipopolysaccharides,
[34] we presumed that incorporation of some specific groups would possibly favor membrane
penetration. Third, in view of the great enhancement of erythromycin antibacterial activity by
carboxy quinolones attached at the 4"-O position of the cladinose, [35-38], systematic evaluation
of various substituted quinolones (n - t, Chart 1) linked to other positions of the aglycone would
be valuable for unveiling the SAR. [39, 40] Therefore, 2-fluoro 9-oxime ketolides 8, 9, 14 and 20
(Ar = a - t, presented in Chart 1) were prepared as illustrated in Schemes 1 - 4.
<
Chart 1>
2
.2 Chemistry
Following a modified procedure, [23] 4 was obtained by diacetylation of the starting material
, and then a propargyl group was regioselectively installed on the 9-oxime of 4 (Scheme 1). The
3
resulting intermediate 5 was converted to 6 by cyclic carbonation at the 11,12-positions and
subsequent oxidation of 3-OH. Next, stereoselective 2-fluorination was completed to afford 7. [41]
Sonogashira coupling with various heteroaryl halide reagents following by deacetylation at the
2
ʹ-position yielded the targets 8 (8a, 8b, 8c, 8e, 8f, 8g, 8h, 8k, 8l, 8n, 8p, 8q, 8r, 8s, 8t).
Interestingly, hydroxy- or carboxy-containing aryl bromides did not work in the Sonogashira
reaction but aryl iodides did react. Hydrogenation of 8p led to 9p with a saturated linker at the
9
-oxime (Scheme 2). Similar to Scheme 1, the target ketolides 14 (14c, 14d, 14g, 14i, 14j, 14m,
1
4n, 14o, 14r, 14t) with propargyl linkers were prepared in six steps from the common
intermediate 4 (Scheme 3). Hydroxy- or carboxy-containing aryl bromides could be used for Heck
coupling reaction, for example with substituents d, i, j, n, and o (Chart 1). Tautomerization exists
in n and o, but the existence of quinolones rather than hydroxyl quinolines in n and o was revealed
6

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1
3
by C NMR, as indicated by a peak at ~175 ppm. In the process of Heck reaction for the synthesis
of 8o, a decarboxylated side-product 8n can be also isolated. The starting material 15 for the target
ketolides 20 was prepared according to the literature [33]. Diacetylation of 15 afforded 16, which
was further converted to 17 by regioselective methylation at the 9-oxime following by cyclic
carbonation at the 11,12-positions (Scheme 4). Next, 18 was obtained by oxidation of the 3-OH.
Similarly, 2-fluoro ketolide 19 was prepared by stereoselective fluorination. Finally, Heck reaction
at the 6-allyl group and subsequent methanolysis at the 2ʹ-position led to the series 20 (20g, 20i,
2
0j, 20n, 20o, 20p, 20q, 20r, 20s, 20t). The carbamoyl quinolone iodide reagents were prepared
as illustrated in the Scheme S1 (see supplementary materials).
Schemes 1-4>
<
2
.3 In vitro antibacterial testing
The antibacterial activities of the samples were assessed against
a
panel of
erythromycin-susceptible and clinical erythromycin-resistant isolates, including Gram-positive
bacteria such as Staphylococcus aureus, Streptococcus pneumoniae and Streptococcus pyogenes,
and Gram-negative pathogens such as Haemophilus influenzae and Moraxella catarrhalis. All
isolates tested were identified by 16S rDNA sequencing. S. aureus ATCC 29213, S. pneumoniae
ATCC 49619 and H. influenzae ATCC49247 were used as controls. All the strains, separately
labeled by corresponding genotypes (erm or mef) and phenotypes, are listed in Table 1.
<
Table 1>
Clarithromycin, azithromycin, telithromycin and ciprofloxacin were tested as the reference
compounds. Antibacterial data are presented in Tables 2 - 3 as the minimal inhibitory
concentration (MIC) and minimal bactericidal activity (MBC). MICs were determined by the
standard two-fold broth dilution method recommended by the Clinical and Laboratory Standards
Institute. [42] The samples from 1-8 fold MIC were transported to culture medium and
subcultured for 24 h in the absence of antibiotics. Then, the lowest concentrations where the
number of colonies was reduced to > 3Log were read as MBCs.
1
0
2
.4 Structure-activity relationships
In the first round, we investigated a variety of heteroaryl groups ranging from a to p as
shown in Chart 1. The antibacterial data of the series 8, 9, 14 and 20 (Ar = a - p) are listed in
7

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Table 2. Most of them are more active than the reference clarithromycin, especially against the
resistant bacteria. The SAR indicated most of the tested compounds presented greatly enhanced
antibacterial activity over 8k (Ar = 4-isoquinolyl) against most of the strains. As 8k can be viewed
as a 2-fluoro version of the previously reported 2, [23] we concluded that the further substitutions
on the aryl groups did have beneficial effects on improving the affinity to the potential binding
sites. Unfortunately, the compounds in the 8, 9 and 14 series are inactive against constitutively
resistant S. pneumoniae with the exception of modestly active 8p, 9p and 14o. In contrast, all of
compounds in the 20 series are highly potent against constitutively resistant S. pneumoniae.
Regarding the series of 9-propargyl oximes 8, the attachment point of amino groups on the
aryl groups has little effect on antibacterial activity (8a vs 8c); electron-withdrawing but less
hydrophilic substituents such as nitro and cyano groups are in general superior to amino groups
(8b vs 8a; 8e vs 8c). As a result, we speculated that compounds 8 interact with the bacterial
ribosomal RNA bases probably via π - π interaction rather than hydrogen bonding. Both the
acetylation of 8c and reverse of the resulting acetamide 8f to the carboxamide 8g did not confer
significant influence on the potency. However, further methylation on the nitrogen atom of the
carboxamide resulted in 2- to 4-fold more potency than unsubstituted carbamate (8h vs 8g). The
macrolide 8h acquired high activity against most of the Gram-positive bacteria and H. influenzae
(only 2-fold less than clarithromycin). However, 8h is still weakly potent against constitutively
resistant S. pneumoniae 07P390 and S. aureus PU20 with MIC values of 32 and 256 µg/mL,
respectively. On the other hand, the bicyclic aryl group 8l, a benzo derivative of 8a, significantly
decreased the activity by 4- to 8-fold less than the monocyclic aryl group 8a, and it is inferior to
the unsubstituted isoquinoline 8k. Compound 8p, a hybrid of macrolides and quinolones,
exhibited activity against the two MRSA strains (MICs: 1 µg/mL vs S. aureus PU32; 32 µg/mL vs
S. aureus PU20). Strikingly, the quinolone 8p showed 8-fold improvement over the methylated
carbamate 8h against constitutive erm-encoded isolates S. pneumoniae 07P390 (MICs: 4 µg/mL),
which implied the importance of the quinolones being the terminal aryl groups. Decarboxylation
of 8p and cyclopropyl deletion led to 8n, which is less active than 8p against S. pneumoniae
encoded mef or constitutive erm genes, S. aureus encoded inducible erm or constitutive erm genes
and H. influenzae, but more effective than 8p against susceptible S. pneumoniae and inducible
8

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erm-encoded S. pyogenes. Among the series 8a - 8p, 8p is the best candidate with a balanced
antibacterial spectrum. Noteworthy, 8p showed bactericidal activity against all the tested strains
irrespective of the genotypes and phenotypes.
Previous SAR studies drawn on the 9-oxime ether ketolides indicated that a flexible sidechain
is detrimental to the potency. [23] However, compound 9p, a hydrogenation product of 8p,
retained a good overall antibacterial spectrum, which suggested that installation of a quinolone at
the end of the sidechains has a favorable impact on the activity.
For the series of 9-allyl oximes 14, the beneficial effects of the substituents on the activity
ranked in the order of carbamoyl groups > amino, hydroxy groups > carboxy groups (14g > 14c,
1
4d > 14i; 14m > 14j). Previous SAR on the 9-oxime ether ketolides disclosed the bicyclic aryl
groups were preferable than monocyclic aryl groups. [22, 23] However, this trend was not
observed when substituents were installed onto the aryl groups (14j vs 14i; 14m vs 14c). Thus, the
substituents indeed have important roles in the antibacterial activity. Compounds 14i, 14j and 14o
possessing carboxy groups had higher MIC values of 32 µg/mL than the others in the 14 series
against H. influenzae, while 14g had a lower MIC value of 8 µg/mL, which reflected the
advantage of carbamoyl groups over carboxylic groups. The carbamoylpyridine 14g is more
effective than the quinolone 14o except for S. pneumoniae 07P390 and S. pyogenes 01-968. The 4-
to 8-fold improvement of the carboxy quinolone 14o over the carboxy quinoline 14j indicated the
importance of the quinolone over the analogous aryl groups. Furthermore, the important function
of substituted quinolones was corroborated by the superiority of 14o over the unsubstituted
quinolone 14n. Due to the incorporation of a carboxy quinolone, 14o was endowed with greatly
enhanced activity with an MIC of 4 µg/mL and an MBC of 16 µg/mL against the constitutively
resistant S. pneumoniae 07P390, which is in agreement with the SAR drawn on the series of
9
-propargyl oximes 8.
For the series of the 6-allyl 20, the bicyclic aryl group 20j is far better to the monocyclic aryl
group 20i. The activity of the pyridyl carboxamide 20g was significantly improved in comparison
to the pyridyl carboxylic acid 20i. Comparison of the quinolone analogs 20n, 20o and 20p
indicated that both introduction of a carboxylic acid at the 3-position of quinolones and
1
installation of a cyclopropyl group at N of the quinolones could enhance the antibacterial activity
9

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against of most of tested strains, but an opposite trend was observed against constitutively
resistant S. pneumoniae 07P390. Interestingly, 20i and 20j sharing an aryl carboxylic acid were
subjected to efflux by S. pneumoniae PU 09. A similar trend was also observed in the series of 14
(
14i and 14j). Generally, the 20 series is more potent than the counterparts in the 14 series (20j vs
4j; 20i vs 14i; 20o vs 14o; 20n vs 14n). However, 14g is generally more potent than 20g, except
it is 64-fold less potent against the constitutively resistant S. pneumoniae 07P390.
Table 2>
1
<
Taken together, the SAR clearly suggested that incorporation of the carbamoyl aryl groups g
and h enables the resulting compounds 8g, 8h and 14g to possess superior and balanced
antibacterial profiles over the other aryl groups such as a-f and i-m, but unfortunately they still
failed to address constitutively resistant S. pneumoniae 07P390. In contrast, the 20 series,
irrespective of the aryl groups, exhibited 64-fold lower MICs against the constitutively resistant S.
pneumoniae 07P390 (MICs: 20, 0.25 - 1 µg/mL vs 8, 9, 14, 4 - >32 µg/mL), which suggested that
the substituted positions of the sidechains attached to the aglycones have great impacts on the
potency. Carboxy quinolones have been utilized extensively as common pharmacophores for
quinolone and the recently reported macrolone antibiotics. [10, 26, 37] Impressively,
incorporation of the carboxy quinolones o and p is capable of minimizing this gap between 8, 14
and 20 to the extent of 8-fold (e.g. 20o vs 14o; 20p vs 8p), which highlights the importance of aryl
groups being quinolones for probing the binding sites within the NPET. To further address the low
potencies conferred by the carboxy quinolones, we decided to incorporate the structural
advantages of g and h into quinolones. Following this reasoning, we designed novel quinolones q,
r, s and t featured with carbamoyl groups instead of the original carboxylic acid at the 3-position
of the quinolones.
The antibacterial data obtained for the series 8, 14 and 20 (Ar = q - t) are listed in Table 3. All
of the compounds 8q - 8t are very active against susceptible S. pneumoniae ATCC49619 (MICs of
≤
0.008 µg/mL). Significantly, it is the first time that the 9-oxime ether ketolides have been
endowed with high activity against constitutive S. pneumoniae (MICs of 8q - 8t and 14q - 14t:
0
.062 - 0.25 µg/mL) and constitutive S. pyogenes (MICs of 8q - 8t and 14q - 14t: 0.125 - 1
1
µg/mL). Among compounds 8q - 8t, the macrolone 8r with an ethyl group on the N position is
1
0

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the most potent, particularly with an MIC of 0.125 µg/mL against constitutively resistant S.
1
pneumoniae 07P390. Compound 8q with a cyclopropyl group on the N position is less active than
8
r. For example, it is 2-fold less potent against constitutive S. pneumoniae 07P390 and 8-fold less
potent against constitutive S. pyogenes. In contrast, 14r is more potent than 8r, featuring an MIC
of 0.062 µg/mL against constitutively resistant S. pneumoniae 07P390, except for H. influenzae
(MBCs: 16 µg/mL of telithromycin, 32 µg/mL of 8r vs 256 µg/mL of 14r) and constitutive S.
pyogenes (MICs: 0.031 µg/mL of telithromycin, 0.125 µg/mL of 8r vs 0.5 µg/mL of 14r).
Generally, an unsubstituted carbamoyl group is favorable for the activity compared to the
methylated carbamoyl group in the series of 8 (8q vs 8s; 8r vs 8t), in 14 (14r vs 14t) and in 20
1
(
20q vs 20s; 20r vs 20t). Notably, 20q, with a cyclopropyl group on the N position, is the most
potent compound in the 20 series, which is different from the SAR in 8.
Table 3>
<
2
.5 In vivo pharmacokinetics properties and CYP inhibition
Telithromycin, 8r, 14r and 20q were formulated in saline solution containing 1% DMF/ 20%
hydroxypropyl-β-cyclodextrin (w/v). [35] In vivo pharmacokinetics (PK) data were obtained in
-3 male SD rats by oral administration of a single dose of 20 mg/Kg. The blood samples were
2
collected at the time points of 0.25, 0.5, 1, 2, 4, 6, 8 and 24 h. The blood samples were centrifuged
at 2500 g for 5 minutes at 4 °C. Quantitative analysis of drug concentrations was performed by
LC-MS/MS. The pharmacokinetic parameters for the tested compounds such as half-life (t ), the
1/2
peak plasma concentration (Cmax), the time to Cmax (Tmax), systematic plasma clearance (CL),
mean residence time (MRT) and the area under curve (AUC) were calculated by WinNonlin
software (version 5.2).
To determine the inhibition potential of 8r, 14r and 20q in human liver microsomes, a CYP
inhibition test was performed. A mixture of the human microsome CYP 3A4, the substrate
(midazolam) and 0.1 M potassium phosphate buffer was warmed at 37 ºC for 5 min. The tested
compounds (or ketoconazole as a positive inhibitor) and 25 mM NADPH solution were added and
the mixture was incubated at 37 ºC for 10 min. The metabolites were detected by HPLC
(ACQUITY UPLC C18, 1.7 µm, 50 x 2.1 mm I.D., Waters; gradient acetonitrile/water as eluents
at a flow rate of 0.5 mL/min) and a triple quadrupole mass spectrometer (API 5500). The peak
1
1

ACCEPTED MANUSCRIPT
area ratio of metabolite/internal standard was determined for each metabolite. The IC50 values
were calculated using commercial software Graphpad Prism (version 7) (Table 4).
The macrolones 8r, 14r and 20q have 2.5 - 3.6 times prolonged half-life and 2.3 - 2.6 longer
mean residence time over telithromycin (Table 4). The macrolone 14r exhibited 1.5-fold higher
clearance but possesses 1.6-fold higher AUC values, which could be attributed to good absorption.
On the contrary, 8r is inferior to telithromycin regarding Cmax and AUC because its clearance is
too high. Compound 20r is superior to telithromycin with the exception of a longer Tmax. Notably,
1
4r and 20r share a more steady plasma concentration for several hours as compared to
telithromycin with a U-turn PK curve (see supplementary materials, Figure S1), and this improved
PK property is very important for this kind of time-dependent antibiotic without a concentration
dependence.
CYP 3A is the key metabolic enzyme for half of the marketed drugs. It was reported that
cethromycin has an IC50 of 2.7 µM for inhibition of CYP 3A4. [43] CYP inhibition results
indicated that 8r, 14r and 20q are not strong inhibitors on CYP 3A4 (IC50 > 5 µM, Table 4).
Among them, only 20q produced lower enzyme inhibition than telithromycin (IC50: 20q, 14.9 µM
vs telithromycin 11.8 µM).
<
Table 4>
2
.6 Molecular modelling
Molecular docking (Figure 5) was performed based on the structure of the
cethromycin-ribosome complex deposited in the Protein Data Bank (PDB ID 1NWX). The crystal
structure of the 50S ribosomal subunit in complex with cethromycin in the reference [20] was
retrieved from 1NWX. The 11-nitrogen and 2-hydrogen were mutated to an 11-oxygen and
2
9
-fluorine, separately. For compounds 8r and 14r, the 9-carbonyl group was converted to a
-oxime. The sidechain at the 9-oxime was docked in the receptor binding site through a covalent
attachment to the oxygen of the 9-oxime. For compounds 20q and 20r, the 9-carbonyl group was
converted to a 9-methyl oxime. The sidechain at the 6-position was docked in the receptor binding
site through a covalent attachment to the oxygen of 6-OMe. Only residues within 30 angstroms of
the cethromycin were kept to simplify the calculation. Each side chain was docked 10 times and
the best docking solution according to the GoldScore fitness function was chosen as the binding
1
2

ACCEPTED MANUSCRIPT
conformation.
The macrolones 8r and 14r orient their sidechains similarly to cethromycin, but extend into a
new pocket formed by A764, C803, C1772 and A802 (Deinococcus radiodurans numbering). The
4
-oxygen of the quinolone ring of 14r hydrogen bonds to the rRNA base C1772. It was reported
that the nitrogen of the quinoline of cethromycin hydrogen bonds to the pentose of C803. [20]
Compounds 20q and 20r orient the sidechains differently from cethromycin, and to a pocket
surrounded by U2588, G2587, C2589, G2560 and C2565. The nitrogen of 3-carbamoyl group on
the quinolone ring of 20q hydrogen bonds to the phosphate of U2588.
<
Figure 5>
3
. Conclusion
We examined the effects of the substituted heteroaryl groups (a - t) on the antibacterial
activities of thirty-six 2-fluoro 9-oxime ketolides, which are structurally different from
telithromycin-like chemicals that interact with A752 within the NPET. Most of them are highly
active against susceptible bacteria and a variety of inducibly erm- or mef-mediated resistant
pathogens. SAR indicated that 9-propargyl ketolides 8 and 9-allyl ketolides 14 are significantly
different from 6-allyl ketolides 20, and molecular modelling suggested that they might have new
binding sites. Incorporation of quinolones, such as n, o, p, q, r, s and t, is critical for antibacterial
activities of the resulting novel macrolones with modest to high potencies against constitutively
resistant S. pneumoniae. Particularly, hybrids of macrolides and carbamoyl quinolones 8r, 14r and
2
0q featured high activities against constitutively erm-mediated S. pneumoniae (MICs: ≤ 0.008 to
0
.125 µg/mL of 8r, 14r and 20q vs 256 µg/mL of azithromycin vs 0.016 µg/mL of telithromycin),
constitutively erm-mediated S. pyogenes (MICs: 0.016 to 0.5 µg/mL of 8r, 14r and 20q vs > 256
µg/mL of azithromycin vs 0.031 µg/mL of telithromycin), and multidrug resistant S. aureus with
high-level resistance against current clinical macrolides, β-lactams and quinolones (MICs: 0.25 to
1
µg/mL of 8r, 14r and 20q vs 32 µg/mL of azithromycin vs 64 µg/mL of ciprofloxacin).
Furthermore, in vivo PK results suggested 8r, 14r and 20q are probably metabolically stable
(
t , and MRT) over telithromycin. Macrolones 14r and 20q, unlike 8r, exhibited improved AUC
1/2
in contrast to telithromycin. Compound 20q is more safe than telithromycin due to its reduced
1
3

ACCEPTED MANUSCRIPT
level of inhibition of human liver CYP 3A4, a key metabolic enzyme responsible for metabolism
of half of the marketed drugs. It is widely acknowledged that additional modes of action are
required for new macrolides to treat constitutively resistant pathogens. This work provides novel
chemotypes for discovery of the new potential binding sites and it also paves the way for
structural insight into unexplored functions of the ribosomal nascent peptide exit tunnel.
4
. Experimental Section
Reagents and solvents were purchased from commercial suppliers, Innochem and J&K
scientific, and were used without further purification. All non-aqueous reactions were carried out
under an atmosphere of argon using dry solvents, unless otherwise noted. All of the reactions were
monitored by thin-layer chromatography (TLC) using silica gel HSGF254 precoated plates (0.2
mm), and the compounds were visualized under UV light (λ = 254 or 365 nm) and/or stained with
iodine. Column chromatography was performed with the indicated eluents and silica gel (100-200
1
13
or 200-300 mesh) purchased from Qingdao Haiyang. H and C spectra were taken in CDCl3,
DMSO-d6 or acetone-d6 on Bruker ARX 400 MHz and 700 MHz spectrometers with
tetramethylsilane (TMS) as an internal standard. The purities of the target compounds for
antibacterial evaluation were examined by TLC under the conditions of different eluents and
visualization, and further confirmed by NMR spectra.
4
.1 2ʹ-O-Acetyl-3-O-descladinosyl-3-hydroxy-6-O-methylerythromycin A 9-O-acetyl oxime (4)
To a solution of 3-O-Descladinosyl-3-hydroxy-6-O-methylerythromycin A 9-O-oxime (3)
(5.433 g, 8.984 mmol) in DCM (50 mL), acetic anhydride (2.600 mL, 26.951 mmol) was added,
and the mixture was kept stirring at room temperature for 1-1.5 h. The mixture was washed with a
saturated NaHCO solution (×5), water, and then brine. The organic layer was concentrated to
3
yield 4 (4.526 g, 6.570 mmol, 73.13%).
4
.2 2ʹ-O-Acetyl-3-O-descladinosyl-3-hydroxy-6-O-methylerythromycin A 9-O-propargyl oxime
11,12- cyclic carbonate (5)
t
KOBu (1.334 g, 11.979 mmol) was added to a solution of compound 4 (5.501 g, 7.986 mmol)
in THF (50 mL) and DMSO (50 mL). The mixture was stirred for 20 min at room temperature.
t
Then KOBu (0.897 g, 7.986 mmol) and 80% propargyl bromide (0.900 mL, 7.986 mmol) were
added to the reaction mixture. After stirring for 1 h, the reaction mixture was poured into H O (50
2
mL) and extracted with EtOAc. The organic layer was washed with water and brine, and
concentrated to yield an intermediate 9-O-propargyl-4 (4.906 g, 7.164 mmol, 89.71%).
1
4

ACCEPTED MANUSCRIPT
To an ice-cold solution of 9-O-propargyl-4 (4.906 g, 7.164 mmol) and pyridine (6.900 mL,
8
5.252 mmol) in DCM (70 mL), a solution of BTC (4.252 g, 14.328 mmol) in DCM (50 mL) was
added dropwise at -10 ºC, and the mixture was stirred for 4 h, after which time the mixture was
increased to room temperature with further stirring for 18 h. Then the temperature was allowed to
cool to 0 ºC, and brine (60 mL) was added dropwise. The organic layer was washed with saturated
NaHCO , water and brine. The mixture was concentrated and purified by column chromatography
3
(
silica gel, DCM/EtOH/NH ·H O = 10:0.05:0.05) to give 5 (3.056 g, 4.299 mmol, 60.00%).
3 2
4
.3 2ʹ-O-Acetyl-3-O-descladinosyl-3-oxo-6-O-methylerythromycin A 9-O-propargyl oxime 11,12-
cyclic carbonate (6)
A solution of NCS (1.654 g, 12.384 mmol) and DMS (1.100 mL, 14.706 mmol) in DCM (20
mL) was stirred for 30 min at -15 ºC. Then a solution of compound 5 (5.502 g, 7.740 mmol) in
DCM (50 mL) was added dropwise at -15 ºC, and the mixture was stirred for 3 h, after which time
the temperature was increased to -5 ºC. Trimethylamine was added to the reaction mixture, and the
mixture was stirred for 1 h. The mixture was allowed to warm to room temperature. The organic
layer was washed with saturated NaHCO , water and brine. The mixture was concentrated to yield
3
6
(5.925 g, 8.359 mmol, 67.50%)
4
.4 2ʹ-O-Acetyl-2-fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
9-O-propargyl
oxime 11,12- cyclic carbonate (7)
An ice-cold solution of 6 (5.925 g, 8.358 mmol) and 60% NaH (0.669 g, 16.716 mmol) in
DMF (20 mL) was stirred for 1 h. NFSI (2.899 g, 9.194 mmol) was added, and the mixture was
further stirred for 3 h. The mixture was allowed to warm to room temperature. The reaction
mixture was poured into H O (50 mL) and extracted with EtOAc. The organic layer was washed
2
with water and brine. The mixture was concentrated and purified by column chromatography
(
silica gel, DCM/NH ·H O = 10: 0.05) to give 7 (2.094 g, 2.881 mmol, 34.47%).
3 2
4
.5 General Procedure for the Synthesis of compounds 8 series.
To a solution of CuI (0.1 eq), Pd(PPh ) Cl (0.05 eq) and compound 7 (1 eq) in MeCN (5 mL)
3
2
2
were added ArBr (Ar=a, b, c, e, f, g, h, k, l) or ArI (Ar=q, r, s, t) (3 eq) and trimethylamine (1.5
eq). The reaction mixture was flushed with argon and sealed in a pressure tube. The reaction
mixture was stirred at 80 ºC for 3-4 h. The reaction mixture was extracted with EtOAc, washed
with water and brine and concentrated in vacuo. The product was dissolved in MeOH (15 mL) at
6
0 ºC for 1 h. The organic solvent was removed in vacuum. The crude mixture was purified by
column chromatography on silica gel to give 8a, 8b, 8c, 8e, 8f, 8g, 8h, 8k, 8l, 8q, 8r, 8s, 8t.
To a solution of ArI (Ar=n, p) (3 eq) in MeCN (5 mL) was added trimethylamine (5 mL).The
mixture was stirred at room temperature for 1 h, after which time CuI (0.1 eq), Pd(PPh ) Cl (0.05
3
2
2
eq) and compound 7 (1 eq) were added. The reaction mixture was flushed with argon and sealed in
a pressure tube. The reaction mixture was stirred at 50 ºC for 24 h. The reaction mixture was
1
5

ACCEPTED MANUSCRIPT
extracted with EtOAc, washed with water and brine and concentrated in vacuum. The product was
then dissolved in MeOH (15 mL) at 60 ºC for 1 h. The organic solvent was removed in vacuo. The
crude mixture was purified by column chromatography on silica gel to give 8n and 8p.
4
9
.5.1
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(2ʹ-aminopyrid-5ʹ-yl)-2-propargyl] oxime 11,12- cyclic carbonate (8a)
According to the general procedure, from compound 7 (0.550 g, 0.800 mmol) and
-amino-5-bromopyridine (0.480 g, 2.800 mmol), 8a (27.0 mg, 0.035 mmol, 4.30%) was obtained
2
as an off-white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:0.1:0.05).
3
2
+
1
HRMS (ESI) (M+H) m/z 777.4073, calcd for C H FN O 777.4081. H NMR (CDCl , 500
3
9
58
4
11
3
MHz) δ: 8.13 (s, 1 H, 6ʹ-Ar), 7.43 (dd, J = 8.5 Hz, J = 2.0 Hz, 1 H, 3ʹ-Ar), 6.41 (d, J = 8.5 Hz, 1
1
2
H, 4ʹ-Ar), 4.96 (d, J = 8.5 Hz, 1 H, H-13), 4.86 (d, J = 15.5 Hz, 1 H, 9-O-CH C≡C-Ar), 4.83 (d, J
2
=
15.5 Hz, 1 H, 9-O-CH C≡C-Ar), 4.32 (d, J = 7.0 Hz, 1 H, H-1ʹ), 4.02 (d, J = 9.5 Hz, 1 H, H-5),
2
3
2
2
6
.58-3.50 (m, 4 H, H-8, H-5ʹ, H-4, 2ʹ-OH), 3.22-3.19 (m, 1 H, H-2ʹ), 2.62 (s, 3 H, 6-O-CH3),
.55-2.51 (m, 2 H, H-3ʹ, H-10), 2.31 (s, 6 H, -N(CH ) ), 1.94-1.91 (m, 1 H, H-14eq), 1.76 (d, J =
3
2
1.0 Hz, 3 H, 2-CH ), 1.68-1.64 (m, 3 H, H-7a, H-14ax, H-4ʹa), 1.59 (s, 3 H, 12-CH ), 1.42 (s, 3 H,
3
3
-CH ), 1.28-1.22 (m, 10 H, 10-CH , 5ʹ-CH , 4-CH , H-7b), 1.00 (s, 3 H, 8-CH ), 0.93 (t, J = 7.5
3
3
3
3
3
Hz, 3 H, 15-CH3).
4
.5.2
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
9
-O-[3-(2ʹ-nitropyrid-5ʹ-yl)-2-propargyl] oxime 11,12- cyclic carbonate (8b)
Following the general procedure, from compound 7 (0.300 g, 0.413 mmol) and
-bromo-2-nitropyridine (0.251 g, 1.239 mmol), 8b (125 mg, 0.155mmol, 37.5%) was obtained as
5
an off-white solid (column chromatography eluents: DCM/NH ·H O = 10:0.05). HRMS (ESI)
3
2
+
1
(
M+H) m/z 807.3845, calcd for C H FN O 807.3822. H NMR (CDCl , 400 MHz) δ: 8.62 (d,
39 56 4 13 3
J = 1.9 Hz, 1 H, 6ʹ-Ar), 8.22 (d, J = 8.3 Hz, 1 H, 3ʹ-Ar), 8.03 (dd, J = 8.4 Hz, J = 2.1 Hz, 1 H,
1
2
4
ʹ-Ar), 4.97 (dd, J = 9.8 Hz, J = 3.0 Hz, 1 H, H-13), 4.91 (s, 2H, 9-O-CH C≡C-Ar), 4.68 (s, 1 H,
1 2 2
H-11), 4.31 (d, J = 7.2 Hz, 1 H, H-1ʹ), 4.05 (d, J = 10.4 Hz, 1 H, H-5), 3.77-3.46 (m, 3 H, H-5ʹ,
H-8, H-4), 3.21 (dd, J = 10.1 Hz, J = 7.2 Hz, 1 H, H-2ʹ), 2.62 (s, 3 H , 6-O-CH ), 2.56-2.45 (m, 2
1
2
3
H, H-3ʹʹ, H-10), 2.30 (s, 6 H, -N(CH ) ), 2.00-1.89 (m, 1H, H-14eq), 1.78 (d, J = 21.4 Hz, 3H,
3
2
2
-CH ), 1.73-1.68 (m, 3 H, H-4ʹa, H-14ax, H-7a), 1.60 (s, 3 H, 12-CH ), 1.42 (s, 3 H, 6-CH ),
3 3 3
1
.33-1.20 (m, 11 H, 5ʹ-CH , H-7b, 4-CH , H-4ʹb, 10-CH ), 1.03 (br, 3 H, 8-CH ), 0.94 (t, J = 7.4
3
3
3
3
1
3
Hz , 3 H, 15-CH ). C NMR (CDCl , 100 MHz) δ:203.1, 202.8, 166.1, 165.8, 155.0, 151.2, 142.3,
3
3
1
2
4
9
26.1, 117.6, 104.2, 94.6, 80.9, 77.5, 77.2, 70.4, 69.5, 65.9, 61.7, 49.1, 40.3, 38.6, 28.3, 25.1, 24.9,
2.4, 21.2, 20.1, 18.8, 15.5, 15.0, 13.1, 10.4.
.5.3
-O-[3-(3ʹ-aminopyrid-5ʹ-yl)-2-propargyl] oxime 11,12- cyclic carbonate (8c)
According to the general procedure, from compound 7 (0.600 g, 0.880 mmol) and
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
1
6

ACCEPTED MANUSCRIPT
3
-amino-5-bromopyridine (0.530 g, 3.080 mmol), 8c (21 mg, 0.027 mmol, 3.10%) was obtained
as a white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:0.1:0.05). HRMS
3
2
+
1
(
ESI) (M+H) m/z 777.4078, calcd for C H FN O 777.4081. H NMR (CDCl , 400 MHz) δ:
39 58 4 11 3
8
.10-7.98 (m, 2 H, 2ʹ-Ar, 6ʹ-Ar), 7.05 (s, 1 H, 4ʹ-Ar), 4.97 (d, J = 8.8 Hz, 1 H, H-13), 4.85 (s, 2 H,
9
-O-CH C≡C-Ar), 4.69 (s, 1 H, H-11), 4.32 (d, J = 6.8 Hz, 1 H, H-1ʹ), 4.03 (d, J = 10.0 Hz, 1 H,
2
H-5), 3.66-3.52 (m, 4 H, H-8, H-5ʹ, H-4, 2ʹ-OH), 3.24 (dd, J = 8.0 Hz, J = 0.4 Hz, 1 H, H-2ʹ),
1
2
2
.62 (s, 3 H, 6-O-CH ), 2.57-2.50 (m, 2 H, H-3ʹ, H-10), 2.35 (s, 6 H, -N(CH ) ), 1.97-1.91 (m, 1 H,
3 3 2
H-14eq), 1.77 (d, J = 21.2 Hz, 3 H, 2-CH ), 1.70-1.64 (m, 3 H, H-7a, H-14ax, H-4ʹa), 1.59 (s, 3 H,
3
1
8
1
6
1
4
9
2-CH ), 1.42 (s, 3 H, 6-CH ), 1.29-1.24 (m, 10 H, 10-CH , 5ʹ-CH , 4-CH , H-7b), 1.02 (br, 3 H,
3 3 3 3 3
1
3
-CH ), 0.93 (t, J = 7.2 Hz, 3 H, 15-CH ). C NMR (CDCl , 176 MHz) δ: 203.0, 202.9, 166.0,
3
3
3
65.9, 165.3, 154.4, 142.3, 136.7, 123.9, 103.9, 98.5, 97.1, 88.3, 84.5, 83.2, 80.9, 78.8, 77.7, 70.2,
9.2, 66.1, 61.9, 49.2, 40.9, 40.3, 38.7, 33.1, 28.8, 26.7, 25.1, 24.9, 22.4, 21.1, 20.1, 18.8, 15.6,
5.0, 13.2, 10.5.
.5.4
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3ʹ-cyanopyrid-5ʹ-yl)-2-propargyl] oxime 11,12- cyclic carbonate (8e)
According to the general procedure, from compound 7 (0.300 g, 0.413 mmol) and
-bromo-3-cyanopyridine (0.227 g, 1.239 mmol), 8e (75 mg, 0.0953 mmol, 23.08%) was obtained
5
as a white solid (column chromatography eluents: petroleum/DCM/EtOH/NH ·H O = 4:6:2:0.1).
3
2
+
1
HRMS (ESI) (M+H) m/z 787.3925, calcd for C H FN O 787.3924. H NMR (CDCl , 400
4
0
55
4
11
3
MHz) δ: 8.88 (d, J = 1.9 Hz, 1 H, 2ʹ-Ar), 8.78 (d, J = 1.9 Hz, 1 H, 6ʹ-Ar), 7.98 (s, 1 H, 4ʹ-Ar), 4.98
(dd, J = 9.8 Hz, J = 3.1 Hz, 1 H, H-13), 4.88 (s, 2 H, 9-O-CH C≡C-Ar), 4.67 (s, 1 H, H-11), 4.30
1 2 2
(
d, J = 7.2 Hz, 1 H, H-1ʹ), 4.04 (d, J = 10.4 Hz, 1 H, H-5), 3.78-3.47 (m, 3 H, H-5ʹ, H-8, H-4),
.19 (dd, J = 10.1 Hz, J = 7.3 Hz, 1 H, H-2ʹ), 2.62 (m, 3 H , 6-O-CH ), 2.52-2.41 (m, 2 H, H-3ʹ,
3
1
2
3
H-10), 2.28 (s, 6 H, -N(CH ) ), 2.00-1.89 (m, 1H, H-14eq), 1.78 (d, J = 21.4 Hz, 3H, 2-CH ),
3
2
3
1
.73-1.62 (m, 3 H, H-4ʹa, H-14ax, H-7a), 1.60 (s, 3 H, 12-CH ), 1.42 (s, 3 H, 6-CH ), 1.33-1.21
3 3
(
m, 11 H, 5ʹ-CH , H-7b, 4-CH , H-4ʹb, 10-CH ), 1.03 (br, 3 H, 8-CH ), 0.94 (t, J = 7.5 Hz , 3 H,
3 3 3 3
1
3
1
5-CH3). C NMR (CDCl , 100 MHz) δ: 203.2, 202.9, 166.0, 165.7, 155.2, 150.8, 141.4, 120.7,
3
1
15.8, 109.8, 104.2, 96.7, 92.2, 80.9, 80.4, 77.5, 77.2, 70.4, 69.6, 65.9, 61.6, 49.1, 40.2, 38.7, 33.1,
8.2, 26.7, 25.2, 24.9, 22.3, 21.2, 20.1, 18.9, 15.6, 15.0, 13.2, 10.5.
.5.5 2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
-O-[3-(3'-acetamidopyrid-5'-yl )-2-propargyl] oxime 11,12- cyclic carbonate (8f)
2
4
9
A
Following the general procedure, from compound 7 (0.300 g, 0.413 mmol) and
-acetamido-5-bromopyridine (0.266 g, 1.239 mmol), 8f (32.5 mg, 0.0397 g, 9.61%) was obtained
3
as a white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:0.5:0.1). HRMS
3
2
+
1
(
ESI) (M+H) m/z 819.4176, calcd for C H FN O 819.4186. H NMR (CDCl , 400 MHz) δ:
41 60 4 12 3
8
.80 (s, 1 H, Ar-NH-CH ), 8.32 (s, 1 H, 6ʹ-Ar), 8.03 (s, 1 H, 4ʹ-Ar), 8.01 (s, 1 H, 2ʹ-H), 5.01 (d, J=
3
1
7

ACCEPTED MANUSCRIPT
9
.5 Hz, 1 H, H-13), 4.84 (s, 2 H, 9-O-CH C≡C-Ar), 4.73 (s, 1 H, H-11), 4.31 (d, J = 7.2 Hz, 1 H,
2
H-1ʹ), 4.04 (d, J = 10.4 Hz, 1 H, H-5), 3.79-3.47 (m, 3 H, H-5ʹ, H-8, H-4), 3.21 (t, J = 8.8 Hz, 1 H,
H-2ʹ), 2.61 (m, 3 H , 6-O-CH ), 2.58-2.42 (m, 2 H, H-3ʹ, H-10), 2.28 (s, 6 H, -N(CH ) ), 2.19 (s, 3
3
3 2
H, Ar-NH-CH ), 2.03-1.90 (m, 1H, H-14eq), 1.77 (d, J = 21.3 Hz, 3H, 2-CH ), 1.72-1.64 (m, 3 H,
3
3
H-4ʹa, H-14ax, H-7a), 1.61 (s, 3 H, 12-CH ), 1.42 (s, 3 H, 6-CH ), 1.33-1.19 (m, 11 H, 5ʹ-CH ,
3
3
3
H-7b, 4-CH , H-4ʹb, 10-CH ), 1.03 (d, J= 6.7 Hz, 3 H, 8-CH ), 0.95 (t, J = 7.5 Hz , 3 H, 15-CH ).
3
3
3
3
1
3
C NMR (CDCl , 100 MHz) δ: 203.1, 202.8, 168.8, 166.1, 165.8, 165.4, 154.6, 146.8, 140.3,
3
1
6
1
4
9
34.7, 129.6, 119.9, 104.1, 98.7, 96.7, 89.3, 84.9, 83.4, 82.7, 81.0, 79.0, 77.6, 77.2, 70.4, 69.5,
5.9, 61.7, 49.2, 40.8, 40.3, 38.7, 29.7, 28.3, 26.8, 25.2, 24.9, 24.4, 22.4, 21.2, 20.5, 18.9, 15.6,
5.0, 13.2, 10.5.
.5.6
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3ʹ-carbamoylpyrid-5ʹ-yl)-2-propargyl] oxime 11,12- cyclic carbonate (8g)
According to the general procedure, from compound 7 (0.570 g, 0.770 mmol) and
-bromonicotinamide (0.480 g, 2.370 mmol), 8g (17.22 mg, 0.021 mmol, 2.70%) was obtained as
5
a white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:0.1:0.1). HRMS (ESI)
3
2
+
1
(
(
(
M+H) m/z 805.40241, calcd for C H FN O 805.40298. H NMR (CDCl , 500 MHz) δ: 9.04
40 58 4 12 3
d, J = 2.0 Hz, 1 H, 6ʹ-Ar), 8.73 (d, J = 1.5 Hz, 1 H, 2ʹ-Ar), 8.21 (t, J = 2.0 Hz, 1 H, 4ʹ-Ar), 4.94
dd, J = 3.0 Hz, J = 9.5 Hz, 1 H, H-13), 4.85 (d, J = 3.5 Hz, 2 H, 9-O-CH C≡C-Ar), 4.73 (s, 1 H,
1
2
2
H-11), 4.31 (d, J = 7.5 Hz, 1 H, H-1ʹ), 4.04 (d, J = 10.5 Hz, 1 H, H-5), 3.64-3.51 (m, 3 H, H-8,
H-5ʹ, H-4), 3.21 (dd, J = 10.0 Hz, J = 7.0 Hz, 1 H, H-2ʹ), 2.61 (s, 3 H, 6-O-CH ), 2.52-2.46 (m, 2
1
2
3
H, H-3ʹ, H-10), 2.30 (s, 6 H, -N(CH ) ), 1.94-1.91 (m, 1 H, H-14eq), 1.75 (d, J = 21.5 Hz, 3 H,
3
2
2
1
4
9
-CH ), 1.60 (s, 3 H, 12-CH ), 1.42 (s, 3 H, 6-CH ), 1.29-1.24 (m, 9 H, 10-CH , 5ʹ-CH , 4-CH ),
3 3 3 3 3 3
.03 (d, J = 6.0 Hz, 3 H, 8-CH ), 0.94 (t, J = 7.5 Hz, 3 H, 15-CH ).
3
3
.5.7
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3'-N-methylcarbomoylpyrid-5'-yl)-2-propargyl] oxime 11,12- cyclic carbonate (8h)
According to the general procedure, from compound 7 (0.750 g, 1.090 mmol) and
-bromo-N-methyl-3-pyridinecarboxamide (0.820 g, 3.820 mmol), 8h (65.0 mg, 0.079 mmol,
.33%) was obtained as a white solid (column chromatography eluents: DCM/EtOH/NH ·H O =
5
7
1
3
2
+
1
0:0.05:0.05). HRMS (ESI) (M+H) m/z 819.4222, calcd for C H FN O 819.4186. H NMR
4
1
60
4
12
(
CDCl , 400 MHz) δ: 9.00 (s, 1 H, 6ʹ-Ar), 8.68 (s, 1 H, 2ʹ-Ar), 8.16 (s, 1 H, 4ʹ-Ar), 7.06 (s, 1 H,
3
-
NH-CH ), 4.95 (d, J = 7.6 Hz, 1 H, H-13), 4.84 (s, 2 H, 9-O-CH C≡C-Ar), 4.72 (s, 1 H, H-11),
3
2
4
2
3
.31 (d, J = 7.2 Hz, 1 H, H-1ʹ), 4.03 (d, J = 10.4 Hz, 1 H, H-5), 3.68-3.51 (m, 4 H, H-8, H-5ʹ, H-4,
ʹ-OH), 3.20 (dd, J = 9.2 Hz, J = 8.0 Hz, 1 H, H-2ʹ), 3.00 (d, J = 4.4 Hz, 3 H, -NH-CH ), 2.60 (s,
1
2
3
H, 6-O-CH ), 2.55-2.44 (m, 2 H, H-3ʹ, H-10), 2.27 (s, 6 H, -N(CH ) ), 1.95-1.90 (m, 1 H,
3
3 2
H-14eq), 1.77 (d, J = 21.6 Hz, 3 H, 2-CH ), 1.69-1.66 (m, 3 H, H-7a, H-14ax, H-4ʹa), 1.61 (s, 3 H,
3
1
2-CH ), 1.42 (s, 3 H, 6-CH ), 1.29-1.23 (m, 10 H, 10-CH , 5ʹ-CH , 4-CH , H-7b), 1.02 (d, J = 5.6
3 3 3 3 3
1
8

ACCEPTED MANUSCRIPT
1
3
Hz, 3 H, 8-CH ), 0.94 (t, J = 7.2 Hz, 3 H, 15-CH ). C NMR (CDCl , 100 MHz) δ: 203.1, 202.8,
3
3
3
1
8
2
4
9
66.0, 165.8, 165.4, 165.3, 154.6, 153.9, 147.9, 137.2, 129.5, 119.7, 104.2, 98.7, 96.6, 90.1, 84.9,
3.2, 82.0, 80.9, 78.9, 70.4, 69.5, 65.8, 61.6, 49.1, 40.2, 38.6, 28.2, 26.9, 25.1, 24.9, 22.4, 21.2,
0.0, 18.8, 15.5, 14.9, 13.1, 10.4.
.5.8
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(4ʹ-isoquinolyl)-2-propargyl] oxime 11,12- cyclic carbonate (8k)
According to the general procedure, from compound 7 (0.663 g, 0.910 mmol) and
4
-bromoisoquinoline (0.570 g, 2.740 mmol), 8k (116 mg, 0.170 mmol, 18.70%) was obtained as a
+
white solid (column chromatography eluents: DCM/NH ·H O = 10:0.1). HRMS (ESI) (M+H)
3
2
1
m/z 812.41240, calcd for C H FN O 812.41281. H NMR (CDCl , 500 MHz) δ: 9.17 (s, 1 H,
43
59
3
11
3
1
7
9
ʹ-Ar), 8.64 (s, 1 H, 3ʹ-Ar), 8.29 (d, J = 8.5 Hz, 1 H, 8ʹ-Ar), 7.97 (d, J = 8.0 Hz, 1 H, 5ʹ-Ar),
.80-7.76 (m, 1 H, 7ʹ-Ar), 7.64 (t, J = 7.5 Hz, 1 H, 6ʹ-Ar), 5.02 (d, J = 2.5 Hz, 2 H,
-O-CH C≡C-Ar), 4.95 (d, J = 7.0 Hz, 1 H, H-13), 4.72 (s, 1 H, H-11), 4.30 (d, J = 7.5 Hz, 1 H,
2
H-1ʹ), 4.03 (d, J = 10.5 Hz, 1 H, H-5), 3.82-3.67 (m, 1 H, H-8), 3.53-3.49 (m, 2 H, H-5ʹ, H-4),
.21 (dd, J = 7.5 Hz, J = 10.0 Hz, 1 H, H-2ʹ), 2.58 (s, 3 H, 6-OCH ), 2.52-2.46 (m, 2 H, H-3ʹ,
3
1
2
3
H-10), 2.30 (s, 6 H, -N(CH ) ), 1.97-1.94 (m, 1 H, H-14eq), 1.75 (d, J = 20.5 Hz, 3 H, 2-CH ),
3
2
3
1
.70-1.67 (m, 3 H, H-7a, H-14ax, H-4ʹa), 1.60 (s, 3 H, 12-CH ), 1.46 (s, 3 H, 6-CH ), 1.30-1.26
3 3
(
m, 7 H, 10-CH , 5ʹ-CH , H-7b), 1,22 (d, J = 6.0 Hz, 3 H, 4-CH ), 1.04 (s, 3 H, 8-CH ), 0.94 (t, J
3 3 3 3
1
3
=
7.5 Hz, 3 H, 15-CH3). C NMR (CDCl , 125 MHz) δ: 203.0, 202.8, 165.9, 165.8, 165.6, 154.2,
3
1
6
1
4
9
52.1, 146.5, 135.8, 131.1, 127.8, 127.7, 125.2, 115.5, 104.1, 93.1, 81.0, 80.9, 78.9, 77.7, 70.4,
9.4, 65.9, 62.1, 49.1, 40.2, 38.7, 28.4, 25.0, 24.8, 22.4, 21.1, 20.0, 18.8, 15.5, 15.1, 14.9, 13.2,
0.5.
.5.9
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(1'-aminoisoquinol-4'-yl)-2-propargyl] oxime 11,12- cyclic carbonate (8l)
Following the general procedure, from compound 7 (0.800 g, 1.170 mmol) and
-amino-4-bromoisoquinoline (0.910 g, 1.0mmol), 8l (29.0 mg, 0.035 mmol, 3.01%) was obtained
1
as an off-white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:0.1:0.1).
3
2
+
1
HRMS (ESI) (M+H) m/z 827.4240, calcd for C H FN O 827.4237. H NMR (CDCl , 400
4
3
60
4
11
3
MHz) δ: 8.20 (d, J = 8.0 Hz, 1 H, 8ʹ-Ar), 8.10 (s, 1 H, 3ʹ-Ar), 7.80 (d, J = 8.4 Hz, 1 H, 5ʹ-Ar), 7.72
t, J = 7.6 Hz, 1 H, 7ʹ-Ar), 7.53 (t, J = 7.2 Hz, 1 H, 6ʹ-Ar), 5.56 (s, 1 H, Ar-NH ), 5.02-4.87 (m, 3
(
2
H, 9-O-CH C≡C-Ar, H-13), 4.71 (s, 1 H, H-11), 4.31 (br, 1 H, H-1ʹ), 4.01 (d, J = 10.0 Hz, 1 H,
2
H-5), 3.65-3.49 (m, 4 H, H-8, H-5ʹ, H-4, 2ʹ-OH), 3.22 (br, 1 H, H-2ʹ), 2.57 (s, 3 H, 6-O-CH3),
2
2
6
.53-2.47 (m, 2 H, H-3ʹ, H-10), 2.32 (s, 6 H, -N(CH ) ), 1.97-1.89 (m, 1 H, H-14eq), 1.75 (d, J =
3 2
1.6 Hz, 3 H, 2-CH ), 1.69-1.67 (m, 3 H, H-7a, H-14ax, H-4ʹa), 1.59 (s, 3 H, 12-CH ), 1.44 (s, 3 H,
3
3
-CH ), 1.29-1.21 (m, 10 H, 10-CH , 5ʹ-CH , 4-CH , H-7b), 1.01 (br, 3 H, 8-CH ), 0.93 (t, J = 7.2
3
3
3
3
3
Hz, 3 H, 15-CH3).
1
9

ACCEPTED MANUSCRIPT
4
9
.5.10
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(1',4'-dihydro-4'-oxo-quinol-6'-yl)-2-propargyl] oxime 11,12- cyclic carbonate (8n)
According to the general procedure, from compound 7 (0.200 g, 0.276 mmol) and
-hydroxyl-6-iodoquinoline (0.150 g, 0.552 mmol), 8n (25.5 mg, 0.0308 mmol, 11.15%) was
4
obtained as a white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:0.5:0.1).
3
2
+
1
HRMS (ESI) (M+H) m/z 828.4063, calcd for C H FN O 828.4077. H NMR (Acetone-d , 400
4
3
59
3
12
6
MHz) δ: 8.32 (s, 1 H, 5ʹ-Ar), 7.88 (t, J = 7.2 Hz, 1 H, 2ʹ-Ar), 7.62 (d, J = 8.6 Hz, 1 H, 8ʹ-Ar), 7.93
t, J = 6.2 Hz, 1 H, 7ʹ-Ar), 6.10 (t, J = 7.3 Hz, 1 H, 3ʹ-Ar), 5.05 (d, J = 9.4 Hz, 1 H, H-13), 4.93 (s,
H, 9-O-CH C≡C-Ar), 4.74 (s, 1 H, H-11), 4.35 (d, J = 7.2 Hz, 1 H, H-1ʹ), 4.04 (d, J = 10.3 Hz, 1
(
2
2
H, H-5), 3.80-3.43 (m, 3 H, H-5ʹ, H-8, H-4), 3.15 (t, J = 8.8 Hz, 1 H, H-2ʹ), 2.79 (br, 1 H, H-10),
.63 (s, 3 H, 6-O-CH ), 3.58-2.48 (m, 1 H, H-3ʹ), 2.27 (s, 6 H, -N(CH ) ), 2.00-1.85 (m, 1H,
2
3
3 2
H-14eq), 1.74 (d, J = 21.3 Hz, 3H, 2-CH ), 1.71-1.68 (m, 3 H, H-4ʹa, H-14ax, H-7a), 1.67 (s, 3 H,
3
1
2-CH ), 1.50 (s, 3 H, 6-CH ), 1.32-1.14 (m, 11 H, 5ʹ-CH , H-7b, 4-CH , H-4ʹb, 10-CH ), 1.08 (d,
3 3 3 3 3
1
3
J = 4.4 Hz, 3 H, 8-CH ), 0.94 (t, J = 7.5 Hz , 3 H, 15-CH ). C NMR (Acetone-d , 176 MHz)
3
3
6
δ:205.3, 202.4, 202.3, 176.5, 166.0, 165.8, 153.7, 140.2, 139.0, 134.1, 129.1, 126.2, 118.6, 117.3,
1
4
4
9
09.7, 104.5, 98.6, 97.5, 85.7, 85.3, 84.4, 82.8, 80.6, 78.8, 77.5, 70.4, 69.2, 65.6, 61.6, 48.7, 40.0,
0.0, 38.3, 33.0, 26.7, 24.2, 24.1, 22.2, 20.6, 19.9, 18.0, 15.1, 14.4, 12.6, 8.9.
.5.11
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3'-carboxy-1'-cyclopropyl-1',4'-dihydro-4'-oxo-quinol-6'-yl)-2-propargyl] oxime 11,12-
cyclic carbonate (8p)
According to the general procedure, from compound 7 (0.200 g, 0.276 mmol) and
1
-cyclopropyl-1, 4-dihydro-6-iodo-4-oxo-3-quinolinecarboxylic acid (0.196 g, 0.552 mmol), 8p
(40.1 mg, 0.0440 mmol, 15.94%) was obtained as a white solid (column chromatography eluents:
+
DCM/EtOH/NH ·H O = 10:1.5:0.1). HRMS (ESI) (M+H) m/z 912.4283, calcd for C H FN O
3
2
47 63
3
14
1
9
12.4289. H NMR (Acetone-d , 400 MHz) δ: 8.81 (s, 1 H, 2ʹ-Ar), 8.47 (s, 1 H, 5ʹ-Ar), 8.35 (d, J
6
=
8.9 Hz, 1 H, 7ʹ-Ar), 7.93 (d, J = 8.8 Hz, 1 H, 8ʹ-Ar), 5.10 (d, J = 9.6 Hz, 1 H, H-13), 4.96 (s, 2 H,
9
-O-CH C≡C-Ar), 4.77 (s, 1 H, H-11), 4.36 (d, J = 7.4 Hz, 1 H, H-1ʹ), 4.04 (d, J = 10.3 Hz, 1 H,
2
H-5), 3.93 (br, 1 H, 1H-cyclopropyl), 3.80-3.43 (m, 4 H, H-5ʹ, H-8, H-4, 2ʹ-OH), 3.16 (t, J = 8.8
Hz, 1 H, H-2ʹ), 2.81 (br, 1 H, H-10), 2.61 (s, 3 H, 6-O-CH ), 3.59-2.50 (m, 1 H, H-3ʹ), 2.28 (s, 6 H,
3
-
N(CH ) ), 2.00-1.86 (m, 1H, H-14eq), 1.74 (d, J = 20.8 Hz, 3H, 2-CH ), 1.68 (s, 3 H, 12-CH ),
3
2
3
3
1
.66-1.53 (m, 3 H, H-4ʹa, H-14ax, H-7a), 1.53-1.43 (m, 5 H, 6-CH , 2 H-cyclopropyl), 1.38-1.17
3
(
m, 13 H, 5ʹ-CH , 2 H-cyclopropyl, H-7b, 4-CH , H-4ʹb, 10-CH ), 1.10 (d, J = 7.0 Hz, 3 H,
3
3
3
1
3
8
2
8
2
-CH ), 0.95 (t, J = 7.2 Hz , 3 H, 15-CH ). C NMR (Acetone-d , 176 MHz) δ:205.3, 202.5,
3
3
6
02.3, 177.9, 166.0, 165.8, 165.4, 153.0, 149.0, 141.1, 135.9, 129.3, 125.8, 118.8, 108.7, 104.5,
8.2, 84.4, 84.1, 82.8, 80.6, 78.8, 77.4, 70.4, 69.2, 65.6, 61.4, 48.7, 41.1, 39.8, 38.3, 35.8, 33.0,
6.7, 24.2, 24.1, 22.2, 20.6, 19.9, 18.0, 15.1, 14.4, 12.6, 9.9, 7.6.
2
0

ACCEPTED MANUSCRIPT
4
9
.5.12
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3'-carbamoyl-1'-cyclopropyl-1',4'-dihydro-4'-oxo-quinoline-6'-yl)-2-propargyl]
oxime
11,12- cyclic carbonate (8q)
According to the general procedure, from compound 7 (0.300 g, 0.413 mmol) and
-cyclopropyl-1,4-dihydro-6-iodo-4-oxo-3-quinolinecarboxamide (30) (0.293 g, 0.826 mmol), 8q
1
(20.4 mg, 0.0224 mmol, 5.43%) was obtained as a white solid (column chromatography eluents:
+
DCM/EtOH/NH ·H O = 10:0.1:0.1). HRMS (ESI) (M+H) m/z 911.4453, calcd for C H FN O
3
2
47 64
4
13
1
9
8
5
4
4
11.4448. H NMR (CDCl , 400 MHz) δ: 9.63 (d, J = 4.9 Hz, 1 H, Ar-NH ), 8.87 (s, 1 H, 2ʹ-Ar),
3
2
.54 (s, 1 H, 5ʹ-Ar), 7.93 (dd, J = 8.8 Hz, J = 2.5 Hz, 1 H, 7ʹ-Ar), 7.75 (d, J = 8.8 Hz, 1 H, 8ʹ-Ar),
1
2
.76 (d, J = 5.0 Hz, 1 H, Ar-NH ), 5.01 (d, J = 9.6 Hz, 1 H, H-13), 4.90 (s, 2 H, 9-O-CH C≡C-Ar),
2
2
.71 (s, 1 H, H-11), 4.33 (d, J = 7.1 Hz, 1 H, H-1ʹ), 4.04 (d, J = 10.4 Hz, 1 H, H-5), 3.82-3.44 (m,
H, H-5ʹ, H-8, H-4, 1H-cyclopropyl), 3.29 (t, J = 8.8 Hz, 1 H, H-2ʹ), 2.74-2.48 (m, 5 H, H-3ʹ,
H-10, 6-O-CH ), 2.41 (s, 6 H, -N(CH ) ), 2.03-1.88 (m, 1H, H-14eq), 1.76 (d, J = 21.3 Hz, 3H,
3
3 2
2
1
1
2
-CH ), 1.71-1.64 (m, 3 H, H-4ʹa, H-14ax, H-7a), 1.60 (s, 1 H, 12-CH ), 1.43 (s, 1 H, 6-CH ),
3
3
3
.40-1.12 (m, 15 H, 2 H-cyclopropyl, 5ʹ-CH , 2 H-cyclopropyl, H-7b, 4-CH , H-4ʹb, 10-CH ),
3
3
3
1
3
.03 (br, 3 H, 8-CH ), 0.75 (t, J = 6.8 Hz , 3 H, 15-CH ). C NMR (CDCl , 100 MHz) δ: 203.2,
3
3
3
02.9, 175.9, 166.5, 166.1, 165.8, 165.3, 154.3, 147.8, 140.4, 135.6, 130.6, 127.4, 120.3, 117.0,
12.0, 103.8, 87.4, 84.6, 80.9, 79.7, 70.2, 69.1, 66.1, 62.0, 49.1, 40.8, 40.3, 38.7, 34.8, 34.8, 34.7,
9.0, 25.1, 24.9, 22.4, 21.1, 20.0, 18.8, 15.6, 15.0, 13.2, 10.5, 8.3.
.5.13 2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
-O-[3-(3'-carbamoyl-1',4'-dihydro-1'-ethyl-4'-oxo-quinol-6'-yl)-2-propargyl] oxime 11,12- cyclic
1
2
4
9
A
carbonate (8r)
According to the general procedure, from compound 7 (0.227 g, 0.312 mmol) and
,4-dihydro-1-ethyl-6-iodo-4-oxo-3-quinolinecarboxamide (31) (0.213 g, 0.624 mmol), 8r (26.7
1
mg, 0.0297 mmol, 9.52%) was obtained as a white solid (column chromatography eluents:
+
DCM/EtOH/NH ·H O = 10:0.1:0.1). HRMS (ESI) (M+H) m/z 899.4447, calcd for C H FN O
3
2
46 64
4
13
1
8
8
1
99.4448. H NMR (CDCl , 400 MHz) δ: 9.68 (d, J = 5.1 Hz, 1 H, Ar-NH ), 8.77 (s, 1 H, 2ʹ-Ar),
3
2
.56 (d, J = 2.0 Hz, 1 H, 5ʹ-Ar), 7.74 (dd, J = 8.8 Hz, J = 2.1 Hz, 1 H, 7ʹ-Ar), 7.48 (d, J = 8.9 Hz,
1
2
H, 8ʹ-Ar), 5.98 (d, J = 5.0 Hz, 1 H, Ar-NH ), 5.01 (dd, J = 9.8 Hz, J = 3.0 Hz, 1 H, H-13), 4.90
2
1
2
(
s, 2 H, 9-O-CH C≡C-Ar), 4.72 (s, 1 H, H-11), 4.38 (d, J = 7.0 Hz, 1 H, H-1ʹ), 4.33 (q, J = 7.3 Hz,
2
1
1
6
3
H, 2 H-ethyl), 4.04 (d, J = 10.3 Hz, 1 H, H-5), 3.79-3.47 (m, 3 H, H-5ʹ, H-8, H-4), 3.42 (dd, J =
1
0.0 Hz, J = 7.1 Hz, 1 H, H-2ʹ), 3.12 (q, J = 7.3 Hz, 1 H, H-10), 3.08-2.99 (m, 1 H, H-3ʹ), 2.62 (s,
2
H, -N(CH ) ), 2.60 (s, 3 H, 6-O-CH ), 2.01-1.88 (m, 2 H, H-14eq, H-4ʹa), 1.76 (d, J = 21.3 Hz,
3
2
3
H, 2-CH ), 1.72-1.63 (m, 2 H, H-14ax, H-7a), 1.61 (s, 3 H, 12-CH ), 1.56 (t, J = 7.2 Hz, 3 H, 3
3
3
H-ethyl), 1.49-1.31 (m, 5 H, 6-CH , H-7b, H-4ʹb), 1.31-1.21 (m, 9 H, 5ʹ-CH , 4-CH , 10-CH ),
3
3
3
3
1
3
1
.04 (br, 3 H, 8-CH ), 0.93 (t, J = 7.4 Hz , 3 H, 15-CH ). C NMR (CDCl , 100 MHz) δ: 203.0,
3 3 3
2
1

ACCEPTED MANUSCRIPT
2
02.8, 175.7, 166.6, 166.0, 165.8, 165.3, 154.4, 147.7, 138.4, 135.7, 131.0, 128.0, 120.0, 116.1,
112.1, 103.3, 98.7, 96.7, 87.3, 84.5, 83.0, 80.7, 78.8, 77.6, 70.0, 38.6, 66.1, 62.0, 49.2, 49.1,49.1,
4
1
4
9
5.8, 40.8, 40.3, 38.6, 33.1, 30.1, 26.7, 25.1, 24.9, 22.3, 21.0, 20.0, 18.8, 15.6, 15.0, 14.6, 13.1,
0.5, 8.6.
.5.14
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3'-N-methylcarbomoyl-1'-cyclopropyl-1',4'-dihydro-4'-oxo-quinol-6'-yl)-2-propargyl]
oxime 11,12- cyclic carbonate (8s)
According to the general procedure, from compound 7 (0.265 g, 0.364 mmol) and
-cyclopropyl-1,4-dihydro-6-iodo-4-oxo-N-methyl-3-quinolinecarboxamide (32) (0.201 g, 0.546
1
mmol), 8s (26.5 mg, 0.0286 mmol, 7.87%) was obtained as a white solid (column chromatography
+
eluents: DCM/EtOH/NH ·H O = 10:0.1:0.1). HRMS (ESI) (M+H) m/z 925.4615, calcd for
3
2
1
C H FN O 925.4605. H NMR (CDCl , 400 MHz) δ: 9.80 (d, J = 5.1 Hz, 1 H, Ar-NH-CH ),
4
8
66
4
13
3
3
8
8
9
.88 (s, 1 H, 2ʹ-Ar), 8.52 (d, J = 2.0 Hz, 1 H, 5ʹ-Ar), 7.93 (d, J = 8.8 Hz, 1 H, 7ʹ-Ar), 7.74 (dd, J =
1
.8 Hz, J = 2.0 Hz, 1 H, 8ʹ-Ar), 5.02 (dd, J = 9.8 Hz, J = 3.0 Hz, 1 H, H-13), 4.90 (s, 2 H,
2
1
2
-O-CH C≡C-Ar), 4.71 (s, 1 H, H-11), 4.37 (d, J = 7.1 Hz, 1 H, H-1ʹ), 4.04 (d, J = 10.4 Hz, 1 H,
2
H-5), 3.77-3.47 (m, 4 H, H-5ʹ, H-8, H-4, 1H-cyclopropyl), 3.42 (t, J = 9.9 Hz, J = 7.1 Hz, 1 H,
1
2
H-2ʹ), 3.00 (d, J = 4.9 Hz, 1 H, Ar-NH-CH ), 2.87-2.65 (m, 1 H, H-3ʹ), 2.62 (s, 6 H, -N(CH ) ),
3
3 2
2
2
6
1
2
.60 (s, 3H, 6-O-CH ), 2.57-2.48 (m, 1 H, H-10), 2.02-1.89 (m, 2 H, H-14eq, H-4ʹa), 1.76 (d, J =
3
1.3 Hz, 3H, 2-CH ), 1.71-1.63 (m, 2 H, H-7a, H-14ax), 1.60 (s, 3 H, 12-CH ), 1.42 (s, 3 H,
3
3
-CH ), 1.39-1.13 (m, 15 H, 2 H-cyclopropyl, 5ʹ-CH , 2 H-cyclopropyl, H-7b, 4-CH , H-4ʹb,
3
3
3
1
3
0-CH ), 1.04 (br, 3 H, 8-CH ), 0.97 (t, J = 7.4 Hz , 3 H, 15-CH ). C NMR (CDCl , 100 MHz) δ:
3
3
3
3
03.0, 202.8, 176.0, 166.0, 165.8, 165.3, 165.2, 154.3, 147.2, 140.3, 135.4, 130.5, 127.3, 120.0,
16.9, 112.3, 103.2, 87.2, 84.7, 80.8, 78.9, 77.7, 70.0, 68.5, 66.3, 62.0, 49.1, 40.8, 40.3, 38.6, 34.7,
0.1, 29.7, 26.7, 25.9, 25.1, 24.9, 22.4, 21.0, 20.1, 18.8, 15.6, 15.0, 13.2, 10.4, 8.3.
.5.15 2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
-O-[3-(3'-N-methylcarbamoyl-1',4'-dihydro-1'-ethyl-4'-oxo-quinol-6'-yl)-2-propargyl]
1
3
4
9
A
oxime
11,12- cyclic carbonate (8t)
Following the general procedure, from compound 7 (0.265 g, 0.364 mmol) and
,4-dihydro-1-ethyl-6-iodo-4-oxo-N-methyl-3-quinolinecarboxamide (33) (0.194 g, 0.546 mmol),
1
8
t (14.5 mg, 0.0159 mmol, 0.0436%) was obtained as a white solid (column chromatography
+
eluents: DCM/EtOH/NH ·H O = 10:0.1:0.1). HRMS (ESI) (M+H) m/z 913.4601, calcd for
3
2
1
C H FN O 913.4605. H NMR (CDCl , 400 MHz) δ: 9.85 (d, J = 5.0 Hz, 1 H, Ar-NH-CH ),
4
7
66
4
13
3
3
8
7
9
.78 (s, 1 H, 2ʹ-Ar), 8.57 (d, J = 2.0 Hz, 1 H, 5ʹ-Ar), 7.74 (dd, J = 8.8 Hz, J = 2.1 Hz, 1 H, 7ʹ-Ar),
1 2
.47 (d, J = 8.9 Hz, 1 H, 8ʹ-Ar), 5.02 (dd, J = 10.0 Hz, J = 3.2 Hz, 1 H, H-13), 4.90 (s, 2 H,
1
2
-O-CH C≡C-Ar), 4.71 (s, 1 H, H-11), 4.42-4.27 (m, 3 H, H-1ʹ, 2 H-ethyl), 4.04 (d, J = 10.8 Hz, 1
2
H, H-5), 3.81-3.46 (m, 4 H, H-5ʹ, H-8, H-4), 3.39 (t, J = 10.2 Hz, J = 7.0 Hz, 1 H, H-2ʹ), 3.01 (d,
1
2
2
2

ACCEPTED MANUSCRIPT
J = 4.9 Hz, 1 H, Ar-NH-CH ), 2.98-2.87 (m, 1 H, H-3ʹ), 2.68-2.51 (m, 10 H, 6-O-CH , -N(CH ) ,
3
3
3 2
H-10), 2.04-1.86 (m, 2 H, H-14eq, H-4ʹa), 1.76 (d, J = 21.5 Hz, 3H, 2-CH ), 1.72-1.63 (m, 2 H,
3
H-7a, H-14ax), 1.61 (s, 3 H, 12-CH ), 1.55 (t, J = 7.2 Hz, 1 H, 3 H-ethyl), 1.47-1.17 (m, 15 H,
3
6
-CH , H-7b, H-7a, H-4ʹb, 5ʹ-CH , 4-CH , 10-CH ), 1.04 (br, 3 H, 8-CH ), 0.94 (t, J = 7.5 Hz , 3
3 3 3 3 3
1
3
H, 15-CH3). C NMR (CDCl , 100 MHz) δ: 203.1, 202.8, 175.9, 166.0, 165.8, 165.4, 165.2,
3
1
6
1
4
9
54.3, 147.1, 138.3, 135.6, 131.0, 128.0, 119.8, 116.0, 112.6, 103.3, 87.2, 84.6, 80.8, 77.7, 70.0,
8.7, 66.4, 62.0, 49.1, 40.8, 40.4, 38.6, 29.8, 25.9, 25.1, 24.9, 22.4, 21.0, 20.0, 19.7, 18.8, 15.6,
5.0, 14.6, 13.2, 10.4.
.6
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3'-carboxy-1'-cyclopropyl-1',4'-dihydro-4'-oxo-quinol-6'-yl)-prop] oxime 11,12- cyclic
carbonate (9p)
0% Pd/C (0.0105 g), HCOOH (0.0350 mL, 0.928 mmol), and HCOONH (0.0292 g, 0.464
1
4
mmol) were added to a solution of 8p (0.105 g, 0.116 mmol) in MeOH (10 mL), which was stirred
at room temperature for 16 h under hydrogen atmosphere. The mixture was filtered and washed
with MeOH. The filtrate was concentrated and purified by column chromatography (silica gel,
DCM/EtOH/NH ·H O = 10:1:0.5) to give 9p (20 mg, 0.0218 mmol, 18.79%). HRMS (ESI)
3
2
+
1
(
M+H) m/z 916.4580, calcd for C H FN O 916.4602. H NMR (CDCl , 500 MHz) δ: 8.86 (s,
47 67 3 14 3
1
6
4
3
H, 2ʹ-Ar), 8.32 (s, 1 H, 5ʹ-Ar), 8.12 (d, J = 8.8 Hz, 1 H, 7ʹ-Ar), 7.84 (d, J = 8.6 Hz, 1 H, 8ʹ-Ar),
.63 (d, J = 15.9 Hz, 1 H, 9-O-CH CH CH -Ar), 4.98 (dd, J = 9.5 Hz, J = 2.0 Hz, 1 H, H-13),
2
2
2
1
2
.73 (s, 1 H, H-11), 4.31 (d, J = 7.3 Hz, 1 H, H-1ʹ), 4.14-3.98 (m, 3 H, 9-O-CH CH CH -Ar, H-5),
2
2
2
.73-3.87 (m, 4 H, H-5ʹ, H-8, H-4, 1 H-cyclopropyl), 3.20 (dd, J = 10.2 Hz, J = 7.3 Hz, 1 H,
1
2
H-2ʹ), 2.90 (t, J = 7.4 Hz, 1 H, H-10), 2.61 (s, 1 H, 6-O-CH ), 2.56-2.39 (m, 4 H, H-3ʹ,
3
9
2
-O-CH CH CH -Ar), 2.27 (s, 6 H, -N(CH ) ), 2.15-2.00 (m, 2 H, 9-O-CH CH CH -Ar),
2 2 2 3 2 2 2 2
.00-1.87 (m, 1 H, H-14eq), 1.78 (d, J = 21.4 Hz, 3H, 2-CH ), 1.72-1.63 (m, 2 H, H-7a, H-4ʹa,
3
H-14ax), 1.60 (s, 3 H, 12-CH ), 1.48-1.34 (m, 5 H, 6-CH , 2 H-cyclopropyl), 1.33-1.11 (m, 13 H,
3
3
5
ʹ-CH , 2 H-cyclopropyl, H-7b, 4-CH , H-4ʹb, 10-CH ), 1.01 (d, J = 6.7 Hz, 3 H, 8-CH ), 0.94 (t, J
3 3 3 3
1
3
=
7.4 Hz , 3 H, 15-CH ). C NMR (CDCl , 126 MHz) δ: 203.2, 202.9, 178.6, 167.2, 166.0, 165.9,
3 3
1
7
2
4
63.6, 154.4, 147.6, 140.8, 139.5, 135.1, 128.5, 126.0, 125.8, 117.5, 108.5, 104.2, 98.5, 96.9, 80.8,
8.9, 77.6, 74.8, 72.1, 70.4, 69.6, 65.8, 49.1, 40.2, 38.6, 35.4, 31.5, 30.7, 29.7, 28.2, 25.1, 24.9,
2.4, 21.2, 20.0, 15.0, 13.0, 10.5, 8.2.
.7 2ʹ-O-Acetyl-3-O-descladinosyl-3-hydroxy-6-O-methylerythromycin A 9-O- allyl oxime (10)
Compound 4 (9.129 g, 13.383 mmol) was dissolved in dry THF (70 mL) and DMSO (70 mL)
t
under an argon atmosphere. KOBu (2.252 g, 20.074 mmol) was added immediately, and the
t
reaction mixture was stirred for 20 min at room temperature. KOBu (1.502 g, 13.383 mmol) and
allyl bromide (1.16 mL, 13.383 mmol) were added to the reaction mixture. After stirring for 1 h,
the reaction mixture was then poured into H O (70 mL) and extracted with EtOAc. The organic
2
2
3

ACCEPTED MANUSCRIPT
layer was washed with water and brine, and concentrated to yield 10 (7.809 g, 11.369 mmol,
8
4.95%).
4
.8 2ʹ-O-Acetyl-3-O-descladinosyl-3-hydroxy-6-O-methylerythromycin A 9-O- allyl oxime 11,12-
cyclic carbonate (11)
To an ice-cold solution of 10 (7.809 g, 11.374 mmol) and pyridine (10.950 mL, 135.351
mmol) in DCM (70 mL), a solution of BTC (6.750 g, 22.748 mmol) in DCM (50 mL) was added
dropwise at -10 ºC, and the mixture was stirred for 4 h, after which time the mixture was increased
to room temperature with further stirring for 18 h. Then the temperature was allowed to cool to 0
ºC, and brine (60 mL) was added dropwise. The organic layer was washed with saturated NaHCO3,
water and brine. The mixture was concentrated and purified by column chromatography (silica gel,
DCM/EtOH/NH ·H O = 10:0.2:0.1) to give 11 (2.964 g, 4.158 mmol, 36.55%).
3
2
4
.9 2ʹ-O-Acetyl-3-O-descladinosyl-3-oxo-6-O-methylerythromycin A 9-O- allyl oxime 11,12- cyclic
carbonate (12)
A solution of NCS (0.249 g, 1.864 mmol) and DMS (0.160 mL, 2.214 mmol) in DCM (10
mL) was stirred for 30 min at -15 ºC. Then a solution of compound 11 (0.830 g, 1.165 mmol) in
DCM (50 mL) was added dropwise at -15 ºC, and the mixture was stirred for 3 h, after which time
the temperature was increased to -5 ºC. The trimethylamine was added to the reaction mixture, and
the mixture was stirred for 1 h. The mixture was allowed to warm to room temperature. The
organic layer was washed with saturated NaHCO , water and brine. The mixture was concentrated
3
to yield 12 (0.864 g, 1.215 mmol, 104.19%).
4
.10 2ʹ-O-Acetyl-2-fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin A 9-O- allyl oxime
11,12- cyclic carbonate (13)
An ice-cold solution of 12 (0.864 g, 1.220 mmol) and 60% NaH (0.0976 g, 2.44 mmol) in
DMF (20 mL) was stirred for 1 h; the NFSI (0.423 g, 1.342 mmol) was added, and the mixture
was further stirred for 3 h. The temperature was allowed to increase to room temperature. The
reaction mixture was poured into H O (20 mL) and extracted with EtOAc. The organic layer was
2
washed with water and brine. The mixture was concentrated and purified by column
chromatography (silica gel, DCM/EtOH/NH ·H O = 10:0.1:0.1) to give 13 (0.695 g, 0.954 mmol,
3
2
7
4
8.20%).
.11 General Procedure for the Synthesis of compounds 14 series
To a solution of compound 13 (1 eq) in MeCN (5 mL) were added ArBr (Ar=c, g, m) or ArI
(
Ar=r, t) (2eq), Pd(OAc) (0.2 eq), P(o-MePh) (0.4 eq) and trimethylamine (3 eq). The reaction
2
3
mixture was flushed with argon and sealed in a pressure tube. The reaction mixture was warmed to
0 ºC for 1 h and stirred at 90 ºC for 24 h. The reaction mixture was extracted with EtOAt, washed
6
with water and brine and concentrated in vacuo. The product was then dissolved in MeOH (15 mL)
at 60 ºC for 1 h. The organic solvent was removed in vacuum. The crude mixture was purified by
2
4

ACCEPTED MANUSCRIPT
column chromatography on silica gel to give 14c, 14g, 14m, 14r, 14t.
To a solution of compound 13 (1 eq) in MeCN (5 mL) were added ArBr (Ar=d, i, j, o) (3 eq),
Pd(OAc) (0.3 eq), P(o-MePh) (0.6 eq) and trimethylamine (excessive amount, 5 mL). The
2
3
reaction mixture was flushed with argon and sealed in a pressure tube. The reaction mixture was
warmed to 60 ºC for 1 h and stirred at 90 ºC for 48 h. The reaction mixture was extracted with
EtOAc, washed with water and brine and concentrated in vacuo. The product wasdissolved in
MeOH (15 mL) at 60 ºC for 1 h. The organic solvent was removed in vacuo. The crude mixture
was purified by column chromatography on silica gel to give 14d, 14i, 14j, 14n, 14o.
4
9
.11.1
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3ʹ-amino-pyrid-5ʹ-yl)-E-prop-2-enyl] oxime 11,12- cyclic carbonate (14c)
Following the general procedure, from compound 13 (1.100 g, 1.510 mmol) and
-amino-5-bromopyridine (0.780 g, 4.530 mmol), 14c (110 mg, 0.140 mmol, 14.75%) was
3
obtained as a white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:0.1:0.1).
3
2
+
1
HRMS (ESI) (M+H) m/z 779.42303, calcd for C H FN O 779.42371. H NMR (CDCl , 500
3
9
60
4
11
3
MHz) δ: 7.99 (d, J = 1.5 Hz, 1 H, 6ʹ-Ar), 7.96 (d, J = 2.5 Hz, 1 H, 2ʹ-Ar), 7.01 (t, J = 2.0 Hz, 1 H,
4
9
9
ʹ-Ar), 6.52 (d, J = 16.0 Hz, 1 H, 9-O-CH CH=CH-Ar), 6.39 (dt, J = 6.0 Hz, J = 16.0 Hz, 1 H,
2 1 2
-O-CH CH=CH-Ar), 4.97 (dd, J = 3.0 Hz, J = 9.0 Hz, 1 H, H-13), 4.67 (m, 3 H, H-11,
2
1
2
-O-CH CH=CH-Ar), 4.32 (d, J = 7.0 Hz, 1 H, H-1ʹ), 4.03 (d, J = 10.0 Hz, 1 H, H-5), 3.70-3.67
2
(m, 1 H, H-8), 3.55-3.51 (m, 2 H, H-5ʹ, H-4), 3.24 (m, 1 H, H-2ʹ), 2.63-2.49 (m, 6 H, 6-OCH3,
H-10, H-3ʹ), 2.35 (s, 6 H, -N(CH ) ), 1.97-1.91 (m, 1 H, H-14eq), 1.77 (d, J = 21.5 Hz, 3 H,
3
2
2
-CH ), 1.67-1.63 (m, 1 H, H-14ax), 1.58 (s, 3 H, 12-CH ), 1.39 (s, 3 H, 6-CH ), 1.29-1.24 (m, 9
3 3 3
H, 10-CH , 4-CH , 5ʹ-CH ), 0.99 (d, J = 7.0 Hz, 3 H, 8-CH ), 0.94 (t, J = 7.5 Hz, 3 H, 15-CH ).
3
3
3
3
3
1
3
C NMR (CDCl , 125 MHz) δ: 203.2, 202.9, 166.1, 165.8, 164.2, 154.3, 142.4, 139.0, 136.8,
3
1
4
4
9
32.7, 129.4, 127.9, 118.1, 104.2, 98.7, 96.7, 84.6, 83.1, 80.9, 78.9, 73.9, 70.4, 69.5, 65.9, 49.1,
0.8, 40.3, 38.7, 33.0, 28.3, 26.6, 25.2, 24.9, 22.9, 21.2, 19.9, 18.9, 15.5, 15.0, 13.2, 10.5.
.11.2
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3ʹ-hydroxyl-pyrid-5ʹ-yl)-E-prop-2-enyl] oxime 11,12- cyclic carbonate (14d)
Following to the general procedure, from compound 13 (0.250 g, 0.343 mmol) and
-hydroxyl-5-bromopyridine (0.179 g, 1.029 mmol), 14d (17.3 mg, 0.0222 mmol, 6.47%) was
3
obtained as a white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:0.4:0.1).
3
2
+
1
HRMS (ESI) (M+H) m/z 780.4076, calcd for C H FN O 780.4077. H NMR (CDCl , 400
3
9
59
3
12
3
MHz) δ: 8.15 (s, 1 H, 6ʹ-Ar), 8.09 (s, 1 H, 2ʹ-Ar), 7.28 (s, 1 H, 4ʹ-Ar), 6.56 (d, J = 16.0 Hz, 1 H,
9
9
4
2
-O-CH CH=CH-Ar), 6.41 (dt, J = 16.2 Hz, J = 5.8 Hz, 1 H, 9-O-CH CH=CH-Ar), 4.96 (d, J =
2 1 2 2
.4 Hz, 1 H, H-13), 4.75-4.63 (m, 3 H, 9-O-CH CH=CH-Ar, H-11), 4.31 (d, J = 7.2 Hz, 1 H, H-1ʹ),
2
.03 (d, J = 10.4 Hz, 1 H, H-5), 3.77-3.46 (m, 1 H, H-5ʹ, H-8, H-4), 3.21 (t, J = 5.6 Hz, 1 H, H-2ʹ),
.60 (s, 3 H, 6-O-CH ), 2.55-2.42 (m, 2 H, H-3ʹ, H-10), 2.28 (s, 6 H, -N(CH ) ), 2.00-1.86 (m, 1 H,
3
3 2
2
5

ACCEPTED MANUSCRIPT
H-14eq), 1.78 (d, J = 21.3 Hz, 3H, 2-CH ), 1.71-1.61 (m, 3 H, H-4ʹa, H-14ax, H-7a), 1.59 (s, 3 H,
3
1
3
1
7
1
4
9
2-CH ), 1.40 (s, 3 H, 6-CH ), 1.34-1.15 (m,11 H, H-7b, 4-CH , H-4ʹb, 10-CH , 5ʹ-CH ), 1.01 (br,
3 3 3 3 3
1
3
H, 8-CH ), 0.94 (t, J = 7.5 Hz, 3 H, 15-CH ). C NMR (CDCl , 100 MHz) δ:203.2, 202.9, 166.0,
3
3
3
65.8, 164.4, 154.5, 154.5, 138.7, 136.4, 134.1, 128.7, 128.6, 120.6, 104.1, 80.8, 79.0, 77.6, 77.4,
3.7, 70.4, 69.5, 65.8, 49.1, 40.8, 38.7, 28.3, 26.7, 25.2, 25.0, 22.4, 21.2, 20.0, 18.8, 15.5, 15.0,
3.2, 10.5.
.11.3
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3ʹ-carbamoyl-pyrid-5ʹ-yl)-E-prop-2-enyl] oxime 11,12- cyclic carbonate (14g)
According to the general procedure, from compound 13 (0.645 g, 0.884 mmol) and
-bromonicotinamide (0.533 g, 2.652 mmol), 14g (25.6 mg, 0.0317 mmol, 3.58%) was obtained
5
as a white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:0.8:0.1). HRMS
3
2
+
1
(
ESI) (M+H) m/z 807.4166, calcd for C H FN O 807.4186. H NMR (CDCl , 400 MHz) δ:
40 60 4 12 3
8
.94 (s, 1 H, 6ʹ-Ar), 8.72 (s, 1 H, 2ʹ-Ar), 8.18 (s, 1 H, 4ʹ-Ar), 6.68 (d, J = 16.1 Hz, 1 H,
-O-CH CH=CH-Ar), 6.54 (dt, J = 16.1 Hz, J = 5.9 Hz, 1 H, 9-O-CH CH=CH-Ar), 6.00 (s, 1 H,
9
2
1
2
2
Ar-NH ), 4.95 (dd, J = 9.7 Hz, J = 3.1 Hz, 1 H, H-13), 4.37-4.26 (m, 3 H, 9-O-CH CH=CH-Ar,
2
1
2
2
H-11), 4.31 (d, J = 7.2 Hz, 1 H, H-1ʹ), 4.04 (d, J = 10.4 Hz, 1 H, H-5), 3.75-3.47 (m, 1 H, H-5ʹ,
H-8, H-4), 3.22 (dd, J = 10.1 Hz, J = 7.3 Hz, 1 H, H-2ʹ), 2.58 (s, 3 H, 6-O-CH ), 2.55-2.45 (m, 2
1
2
3
H, H-3ʹ, H-10), 2.30 (s, 6 H, -N(CH ) ), 2.00-1.87 (m, 1 H, H-14eq), 1.78 (d, J = 21.3 Hz, 3H,
3
2
2
-CH ), 1.73-1.61 (m, 3 H, H-4ʹa, H-14ax, H-7a), 1.59 (s, 3 H, 12-CH ), 1.40 (s, 3 H, 6-CH ),
3 3 3
1
.32-1.15 (m,11 H, H-7b, 4-CH , H-4ʹb, 10-CH , 5ʹ-CH ), 1.02 (br, 3 H, 8-CH ), 0.94 (t, J = 7.5
3
3
3
3
1
3
Hz, 3 H, 15-CH ). C NMR (CDCl , 100 MHz) δ: 203.2, 202.9, 167.4, 166.0, 165.8, 164.2, 154.5,
3
3
1
7
1
4
9
50.6, 147.5, 132.6, 132.4, 129.5, 129.0, 128.9, 104.1, 98.7, 96.6, 84.9, 83.2, 80.9, 78.9, 77.4, 73.4,
0.3, 69.5, 65.9, 49.1, 40.8, 40.3, 38.7, 28.4, 26.7, 25.2, 25.0, 22.4, 21.2, 20.0, 18.8, 15.6, 15.0,
3.1, 10.5.
.11.4
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3'-carboxy-pyrid-5'-yl)-E-prop-2-enyl] oxime 11,12- cyclic carbonate (14i)
According to the general procedure, from compound 13 (0.550 g, 0.755 mmol) and
-bromo-3-pyridincarboxylic acid (0.458 g, 2.265 mmol), 14i (27.6 mg, 0.0342 mmol, 4.53%)
5
was obtained as a white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:2:0.5).
3
2
+
1
HRMS (ESI) (M+H) m/z 808.4021, calcd for C H FN O 808.4026. H NMR (CDCl , 400
4
0
59
3
13
3
MHz) δ: 9.09 (s, 1 H, 6ʹ-Ar), 8.66 (s, 1 H, 2ʹ-Ar), 8.35 (s, 1 H, 4ʹ-Ar), 6.65 (d, J = 16.0 Hz, 1 H,
9
9
4
-O-CH CH=CH-Ar), 6.51 (dt, J = 16.1 Hz, J = 5.7 Hz, 1 H, 9-O-CH CH=CH-Ar), 4.96 (d, J =
2 1 2 2
.5 Hz, 1 H, H-13), 4.80-4.62 (m, 3 H, 9-O-CH CH=CH-Ar, H-11), 4.43 (d, J = 7.4 Hz, 1 H, H-1ʹ),
2
.05 (d, J = 10.2 Hz, 1 H, H-5), 3.76-3.44 (m, 4 H, H-5ʹ, H-8, H-4, 2ʹ-OH), 3.18 (d, J = 12.2 Hz, 1
H, H-2ʹ), 2.72 (s, 6 H, -N(CH ) ), 2.68-2.45 (m, 5 H, 6-O-CH , H-3ʹ, H-10), 2.00-1.86 (m, 2 H,
3
2
3
H-14eq, H-4ʹa), 1.77 (d, J = 21.4 Hz, 3H, 2-CH ), 1.72-1.63 (m, 2 H, H-14ax, H-7a), 1.60 (s, 3 H,
3
2
6

ACCEPTED MANUSCRIPT
1
2-CH ), 1.47-1.16 (m, 14 H, 6-CH , H-7b, 4-CH , H-4ʹb, 10-CH , 5ʹ-CH ), 1.03 (d, J = 6.7 Hz, 3
3
3
3
3
3
1
3
H, 8-CH ), 0.93 (t, J = 7.5 Hz, 3 H, 15-CH ). C NMR (CDCl , 100 MHz) δ:203.1, 202.8, 171.0,
3
3
3
1
7
1
4
9
66.0, 165.8, 164.4, 154.2, 149.7, 149.6, 134.3, 132.2, 130.5, 128.9, 128.3, 103.4, 84.5, 83.1, 78.8,
7.3, 73.8, 69.7, 68.2, 66.6, 49.2, 39.8, 38.6, 30.2, 29.7, 25.1, 24.9, 22.4, 21.0, 19.9, 18.9, 15.5,
5.1, 13.2, 10.5.
.11.5
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(3'-carboxy-quinol-6'-yl)-E-prop-2-enyl] oxime 11,12- cyclic carbonate (14j)
According to the general procedure, from compound 13 (0.250 g, 0.343 mmol) and
-bromo-3-quinolinecarboxylic acid (0.259 g, 1.029 mmol), 14j (11.6 mg, 0.0135g, 3.93%) was
6
obtained as a white solid (column chromatography eluents: DCM/EtOH/NH ·H O = 10:2.5:0.1).
3
2
+
1
HRMS (ESI) (M+H) m/z 858.4167, calcd for C H FN O 858.4183. H NMR (DMSO-d , 400
4
4
61
3
13
6
MHz) δ: 9.28 (d, J = 7.3 Hz, 1 H, 2ʹ-Ar), 8.61 (d, J = 6.6 Hz, 1 H, 4ʹ-Ar), 7.94 (s, 1 H, 5ʹ-Ar), 7.92
s, 1 H, 7ʹ-Ar), 7.86 (d, J = 9.2 Hz, 1 H, 8ʹ-Ar), 6.84 (d, J = 16.2 Hz, 1 H, 9-O-CH CH=CH-Ar),
(
2
6
4
.57 (dt, J = 16.1 Hz, J = 5.9 Hz, 1 H, 9-O-CH CH=CH-Ar), 4.89 (d, J =11.4 Hz, 1 H, H-13),
1 2 2
.77-4.63 (m, 2 H, 9-O-CH CH=CH-Ar), 4.58 (s, 1 H, H-11), 4.22 (d, J = 7.2 Hz, 1 H, H-1ʹ), 3.89
2
(
d, J = 10.3 Hz, 1 H, H-5), 3.67-3.46 (m, 3 H, H-5ʹ, H-8, H-4), 3.06 (dd, J = 10.1 Hz, J = 7.3 Hz,
1 2
1
1
H, H-2ʹ), 2.74-2.60 (m, 2 H, H-3ʹ, H-10), 2.50 (s, 3 H, 6-O-CH ), 2.19 (s, 6 H, -N(CH ) ),
3 3 2
.88-1.77 (m, 1 H, H-14eq), 1.72 (d, J = 21.6 Hz, 3H, 2-CH ), 1.66-1.60 (m, 3 H, H-4ʹa, H-14ax,
3
H-7a), 1.58 (s, 3 H, 12-CH ), 1.36 (s, 1 H, 6-CH ), 1.28-1.06 (m,11 H, H-7b, 4-CH , H-4ʹb,
3
3
3
1
4
9
0-CH , 5ʹ-CH ), 1.02 (d, J = 6.4 Hz, 3 H, 8-CH ), 0.85 (t, J = 7.4 Hz, 3 H, 15-CH ).
3
3
3
3
.11.6
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
-O-[3-(4'-amino-quinol-3'-yl)-E-prop-2-enyl] oxime 11,12- cyclic carbonate (14m)
Following the general procedure, from compound 13 (0.580 g, 0.796 mmol) and
-amino-3-bromoquinoline (0.355 g, 1.592 mmol), 14m (22.9 mg, 0.0276 mmol, 3.47%) was
4
obtained as an off-white solid (column chromatography eluents: PE:DCM/EtOH/NH ·H O =
3
2
+
1
3
:7:0.5:0.1). HRMS (ESI) (M+H) m/z 829.4425, calcd for C H FN O 829.4394. H NMR
43 62 4 11
(
CDCl , 400 MHz) δ: 8.56 (s, 1 H, 2ʹ-Ar), 7.96 (d, J = 8.4 Hz, 1 H, 5ʹ-Ar), 7.81 (d, J = 8.4 Hz, 1 H,
3
8
9
ʹ-Ar), 7.61 (d, J = 7.5 Hz, 1 H, 6ʹ-Ar), 7.44 (d, J = 7.7 Hz, 1 H, 7ʹ-Ar), 6.74 (d, J = 15.8 Hz, 1 H,
-O-CH CH=CH-Ar), 6.25 (dt, J = 16.9 Hz, J = 6.0 Hz, 1 H, 9-O-CH CH=CH-Ar), 5.25 (s, 2 H,
2
1
2
2
Ar-NH ), 4.98 (d, J = 9.5 Hz, 1 H, H-13), 4.81-4.60 (m, 3 H, H-11, 9-O-CH CH=CH-Ar), 4.31 (d,
2
2
J = 8.2 Hz, 1 H, H-1ʹ), 4.02 (d, J = 10.5 Hz, 1 H, H-5), 3.76-3.43 (m, 3 H, H-5ʹ, H-8, H-4),
.26-3.14 (m, 1 H, H-2ʹ), 2.56 (s, 3 H, 6-O-CH ), 2.51-2.40 (m, 2 H, H-3ʹ, H-10), 2.26 (s, 6 H,
3
3
-
N(CH ) ), 2.01-1.87 (m, 1 H, H-14eq), 1.76 (d, J = 21.3 Hz, 3H, 2-CH ), 1.70-1.63 (m, 3 H,
3
2
3
H-4ʹa, H-14ax, H-7a), 1.61 (s, 3 H, 12-CH ), 1.41 (s, 3 H, 6-CH ), 1.34-1.16 (m, 11 H, H-7b,
3
3
1
3
4
-CH , H-4ʹb, 10-CH , 5ʹ-CH ), 1.01 (br, 3 H, 8-CH ), 0.95 (t, J = 7.5 Hz, 3 H, 15-CH ). C NMR
3 3 3 3 3
(
CDCl , 100 MHz) δ: 203.0, 202.8, 169.7, 166.0, 165.8, 164.2, 154.6, 149.3, 149.3, 154.6, 149.3,
3
2
7

ACCEPTED MANUSCRIPT
1
6
4
9
47.6, 146.8, 129.4, 129.2, 120.4, 112.0, 111.9, 104.1, 80.4, 78.9, 77.5, 77.2, 74.2, 73.9, 70.4, 69.5,
6.6, 49.9, 49.1, 40.3, 38.7, 28.2, 26.6, 25.1, 25.0, 23.3, 21.2, 18.6, 16.0, 15.0, 14.8, 13.1, 10.4.
.11.7
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
oxime
A
-O-[3-(3'-carboxy-1',4'-dihydro-4'-oxo-quino-6'-yl)-E-prop-2-enyl]
11,12-
cyclic
carbonate (14o)
According to the general procedure, from compound 13 (0.250 g, 0.343 mmol) and
-bromo-4-hydroxyl-3-quinolinecarboxylic acid (0.276 g, 1.029 mmol), 14o (14.9 mg, 0.0170 g,
6
4
.96%) was obtained as an off-white solid (column chromatography eluents:
+
DCM/EtOH/NH ·H O = 10:3:0.1). HRMS (ESI) (M+H) m/z 874.4131, calcd for C H FN O
3
2
44 61
3
14
1
8
8
9
8
74.4132. H NMR (DMSO-d , 400 MHz) δ: 8.72 (s, 1 H, 2ʹ-Ar), 8.08 (s, 1 H, 5ʹ-Ar), 7.71 (d, J =
6
.8 Hz, 1 H, 7ʹ-Ar), 7.62 (d, J = 8.2 Hz, 1 H, 8ʹ-Ar), 6.80 (d, J = 16.1 Hz, 1 H,
-O-CH CH=CH-Ar), 6.43 (dt, J = 16.1 Hz, J = 6.0 Hz, 1 H, 9-O-CH CH=CH-Ar), 4.91 (d, J =
2
1
2
2
.4 Hz, 1 H, H-13), 4.69 (d, J = 5.8 Hz, 1 H, 9-O-CH CH=CH-Ar), 4.58 (s, 1 H, H-11), 4.22 (d, J
2
=
6.8 Hz, 1 H, H-1ʹ), 3.90 (d, J = 10.6 Hz, 1 H, H-5), 3.70-3.45 (m, 1 H, H-5ʹ, H-8, H-4),
3
.12-3.02 (m, 1 H, H-2ʹ), 2.72-2.55 (m, 2 H, H-3ʹ, H-10), 2.48 (s, 3 H, 6-O-CH ), 2.20 (s, 6 H,
3
-
N(CH ) ), 1.90-1.79 (m, 1 H, H-14eq), 1.72 (d, J = 21.8 Hz, 3H, 2-CH ), 1.67-1.61 (m, 3 H,
3
2
3
H-4ʹa, H-14ax, H-7a), 1.58 (s, 3 H, 12-CH ), 1.36 (s, 3 H, 6-CH ), 1.28-1.05 (m, 11 H, H-7b,
3
3
4
4
9
-CH , H-4ʹb, 10-CH , 5ʹ-CH ), 1.01 (br, 3 H, 8-CH ), 0.86 (t, J = 7.4 Hz, 3 H, 15-CH ).
3 3 3 3 3
.11.8
-O-[3-(1',4'-dihydro-4'-oxo-quinol-6'-yl)-E-prop-2-enyl] oxime 11,12- cyclic carbonate (14n)
A descarboxylic acid by-product 14n (11.5 mg, 0.0138 mmol, 4.02%) obtained during
preparation of 14o was purified by column chromatography (DCM/EtOH/NH ·H O = 10:0.3:0.1).
2-Fluoro-3-O-descladinosyl-3-oxo-6-O-methylerythromycin
A
3
2
Compound 14n was also obtained by the Heck coupling reaction of compound 13 and
+
4
8
7
9
7
9
-hydroxy-6-bromoquinoline. HRMS (ESI) (M+H) m/z 830.4224, calcd for C H FN O
4
3
61
3
12
1
30.4234. H NMR (CDCl , 400 MHz) δ: 8.28 (s, 1 H, 5ʹ-Ar), 7.72 (d, J = 7.3 Hz, 1 H, 2ʹ-Ar),
3
.64 (d, J = 8.7 Hz, 1 H, 7ʹ-Ar), 7.46 (d, J = 8.6 Hz, 1 H, 8ʹ-Ar), 6.68 (d, J = 15.9 Hz, 1 H,
-O-CH CH=CH-Ar), 6.42 (dt, J = 16.1 Hz, J = 6.1 Hz, 1 H, 9-O-CH CH=CH-Ar), 6.28 (d, J =
2
1
2
2
.3 Hz, 1 H, 3ʹ-Ar), 4.98 (d, J = 8.1 Hz, 1 H, H-13), 4.71 (s, 1 H, H-11), 4.67 (d, J = 6.1 Hz, 1 H,
-O-CH CH=CH-Ar), 4.29 (d, J = 7.2 Hz, 1 H, H-1ʹ), 4.02 (d, J = 10.4 Hz, 1 H, H-5), 3.80-3.43
2
(
m, 3 H, H-5ʹ, H-8, H-4), 3.19 (dd, J = 10.2 Hz, J = 7.2 Hz, 1 H, H-2ʹ), 2.57 (s, 3 H, 6-O-CH ),
1
2
3
2
2
6
.53-2.38 (m, 2 H, H-3ʹ, H-10), 2.25 (s, 6 H, -N(CH ) ), 2.01-1.87 (m, 1 H, H-14eq), 1.78 (d, J =
3 2
1.3 Hz, 3H, 2-CH ), 1.72-1.61 (m, 3 H, H-4ʹa, H-14ax, H-7a), 1.59 (s, 3 H, 12-CH ), 1.40 (s, 1 H,
3
3
-CH ), 1.35-1.13 (m, 11 H, H-7b, 4-CH , H-4ʹb, 10-CH , 5ʹ-CH ), 1.00 (br, 3 H, 8-CH ), 0.94 (t,
3
3
3
3
3
13
J = 7.4 Hz, 3 H, 15-CH ). C NMR (CDCl , 100 MHz) δ: 203.4, 203.1, 179.0, 166.2, 166.0, 164.2,
3
3
1
39.5, 138.7, 132.8, 131.8, 129.7, 126.3, 126.2, 123.8, 118.6, 109.6, 104.2, 80.8, 79.0, 77.6, 77.2,
4.1, 70.4, 69.6, 65.8, 40.1, 40.8, 40.2, 38.7, 28.1, 25.2, 25.0, 22.4, 21.2, 20.0, 18.8, 15.6, 15.0,
7
2
8