Design and Synthesis of Novel Amino-triazine Analogues as Selective Bruton's Tyrosine Kinase Inhibitors for Treatment of Rheumatoid Arthritis

Article abstract of DOI:10.1021/acs.jmedchem.8b01147

Bruton's tyrosine kinase (BTK) is a promising drug target for the treatment of multiple diseases, such as B-cell malignances, asthma, and rheumatoid arthritis. A series of novel aminotriazines were identified as highly selective inhibitors of BTK by a scaffold-hopping approach. Subsequent SAR studies of this series using two conformationally different BTK proteins, an activated form of BTK and an unactivated form of BTK, led to the discovery of a highly selective BTK inhibitor, 4b. With significant efficacy in models in vivo and good ADME and safety profiles, 4b was advanced into preclinical studies.

Full text of DOI:10.1021/acs.jmedchem.8b01147

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Article
Design and Synthesis of Novel Amino-triazine Analogs as Selective
Bruton’s Tyrosine Kinase Inhibitors for Treatment of Rheumatoid Arthritis
Wataru Kawahata, Tokiko Asami, Takao Kiyoi, Takayuki Irie, Haruka Taniguchi,
Yuko Asamitsu, Tomoko Inoue, Takahiro Miyake, and Masaaki Sawa
J. Med. Chem., Just Accepted Manuscript • Publication Date (Web): 14 Sep 2018
Downloaded from http://pubs.acs.org on September 15, 2018
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Design and Synthesis of Novel Aminoꢀtriazine
Analogs as Selective Bruton’s Tyrosine Kinase
Inhibitors for Treatment of Rheumatoid Arthritis
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Wataru Kawahata,* Tokiko Asami, Takao Kiyoi, Takayuki Irie, Haruka Taniguchi, Yuko
Asamitsu, Tomoko Inoue, Takahiro Miyake and Masaaki Sawa
Research and Development, Carna Biosciences, Inc., 3rd Floor, BMA, 1ꢀ5ꢀ5 Minatojimaꢀ
Minamimachi, Chuoꢀku, Kobe 650ꢀ0047, Japan
KEYWORDS. Kinase inhibitor, Bruton's tyrosine kinase, BTK, CIA model, Rheumatoid
arthritis, Aminoꢀtriazine.
ABSTRACT
Bruton's tyrosine kinase (BTK) is a promising drug target for the treatment of multiple diseases
such as B cell malignances, asthma, and rheumatoid arthritis. A series of novel aminotriazines
were identified as highly selective inhibitors of BTK by a scaffold hopping approach.
Subsequent SAR studies of this series using two conformationally different BTK proteins, an
activated form of BTK and an unactivated form of BTK, led to the discovery of a highly
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selective BTK inhibitor 4b. With significant efficacies in vivo models and a good ADME and
safety profile, 4b was advanced into preclinical studies.
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INTRODUCTION
Bruton's tyrosine kinase (BTK) is a member of the Tec family of nonꢀreceptor tyrosine
kinases, and expressed mainly in all hematopoietic cells, especially in Bꢀcells and myeloid cells,
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but very low expression level of BTK is found in Tꢀcells, natural killer cells and plasma cells.
BTK is essential for Bꢀcell maturation by mediating the Bꢀcell receptor (BCR) signaling, and
also plays a crucial role in macrophages and mast cell activation via the highꢀaffinity IgE
4ꢀ7
receptor (FcεRI) to release several proꢀinflammatory mediators. The implication of BTK in a
number of pathological processes has been reported, and thus BTK has emerged as a promising
target for new therapeutic interventions in a wide array of diseases involving Bꢀcell and/or
macrophage activation, such as Bꢀcell malignancies, asthma, rheumatoid arthritis and systemic
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lupus erythematosus.
Ibrutinib, the first FDAꢀapproved covalent BTK inhibitor, has
demonstrated impressive response rates in patients with mantle cell lymphoma and chronic
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lymphocytic leukemia, and several second generation covalent BTK inhibitors are currently
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being evaluated in clinical trials for treating those Bꢀcell malignancies. These BTK inhibitors
covalently bind to Cys481 residue in the ATP binding site of BTK to inhibit BTK enzymatic
activity irreversibly. However, there is still a high demand for nonꢀcovalent BTK inhibitors for
the treatment of autoimmune diseases because of the propensity of Michael acceptors to form
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reactive metabolites. In addition, the use of kinase inhibitors for nonꢀoncology indications is
thought to be a difficult challenge due to anticipated adverse effects related to offꢀtarget activities
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of inhibitors.
Hence, it is significantly important to optimize kinase selectivity for producing
safer drugs, and a number of selective nonꢀcovalent BTK inhibitors are also under evaluation in a
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clinical trials. As a part of our effort to develop selective and nonꢀcovalent BTK inhibitors, we
previously reported a series of novel pyrimidine analogs as highly selective and reversible
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inhibitors of BTK represented by an initial lead compound 1. Unfortunately, efforts to optimize
the compound 1 were unsuccessful due to the hERG inhibitory activities of this chemical series,
which would cause QT prolongation. In this article, we describe the design and synthesis of
novel and potent triazineꢀbased inhibitors for BTK to overcome the hERG issue by a scaffold
hopping approach from compound 1.
RESULTS AND DISCUSSION
In the course of our work on the pyrimidineꢀbased analogs, those compounds were found to
show hERG inhibitions, which seemed difficult to eliminate by modifying substituents.
Therefore, we turned our attention to exploration of a new scaffold. Namely, we got interested in
replacing the pyrimidine core with a Nꢀcontaining heteroaryl group, in which the nitrogen atom
would act as the Hꢀbond acceptor to interact with the hinge region of BTK (Figure 1).
Firstly, we synthesized heteroaryl analogs 2aꢀ2c by replacing the pyrimidine ring of compound
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with other possible Nꢀcontaining heteroaryl rings, and tested for the inhibition of two
conformationally different recombinant human BTK enzymes, an activated conformation of
BTK (BTK[A]) and an unactivated conformation of BTK (BTK[U]), based on the fact that
compounds preferentially binding to BTK[U] would show higher kinase selectivity than
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compounds bind to BTK[A] as we previously reported.
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It can be seen from the results in Table 1, all compounds synthesized here exhibited more
potent inhibition for BTK[U] as expected, because the parent compound 1 was designed to bind
BTK[U] preferentially. The pyrazine (2a), 1,3,4ꢀtriazine (2b) and 1,3,5ꢀtriazine (2c) showed
weak inhibition for BTK[A] with IC values in the micromolar range (1.8 ꢀ5.6 µM). The 1,3,4ꢀ
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triazine (2b) and 1,3,5ꢀtriazine (2c) displayed strong inhibition for BTK[U] (IC = 10 and 6.4
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nM, respectively), but the pyrazine (2a) was less potent for BTK[U] (IC = 100 nM). These
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results suggested that the nitrogen atom at 3ꢀposition of the central heteroaryl ring was important
for the inhibitory potency for BTK[U]. In our previous paper, the docking model of the
pyrimidine analog with BTK suggested that the 4ꢀposition of the pyrimidine ring seem to be able
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to interact with the hinge region of BTK. Therefore, we next examined the effects of
substituents at the 4ꢀposition of the 1,3,5ꢀtriazine ring. Introduction of methyl group resulted in a
decrease of inhibitory potency (2d, IC = 7800 and 37 nM for BTK[A] and [U], respectively).
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Incorporation of amino group into compound 2c to give compound 2e, which displayed a
dramatic improvement of inhibitory potency for both BTK[A] and [U] with IC values of 190
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and 0.67 nM, respectively. The methoxy analog 2f was significantly less potent for BTK[U]
IC = 80 nM), and the demethylation (2g) led to further decrease of the inhibitory potency even
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though it has a hydrogen bond donor similar to the amino analog (IC = 8800 nM for BTK[U]).
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To probe the binding mode of the 1,3,5ꢀtriazine analog, we determined the Xꢀray coꢀcrystal
structure of 2e complexed with BTK. As shown in Figure 2, 2e binds in the ATP binding pocket
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of BTK adopting a Srcꢀlike inactive conformation which has been reported previously. The tꢀ
butylphenyl group of compound 2e occupies a back pocket, soꢀcalled the H3 pocket. Compound
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e also forms several important hydrogen bonds with the hinge region of BTK. The NH at 2ꢀ
position and the nitrogen atom at 3ꢀposition of the central triazine ring make two hydrogen bonds
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with the backbone carbonyl and NH of Met477, respectively. The NH at 4ꢀposition of the
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triazine ring is engaged in an extra Hꢀbond interaction with the backbone carbonyl of Glu475.
The benzene ring at 6ꢀposition of the triazine makes a favorable hydrophobic interaction with
Val416, and the methyl substituent on the benzene further enhances this hydrophobic interaction.
The Xꢀray coꢀcrystal structure of 2e bound to BTK revealed that more space is available around
this methyl group, where Asp539 and Lys430 can provide additional interactions with the
compound. Thus, we synthesized 2h by introducing a hydroxyl group at the methylbenzene
which is capable of making Hꢀbonds. As expected, this modification further improved the
inhibitory potency for both BTK[A] and BTK[U] (Table 1, IC = 43 and <0.3 nM for BTK[A]
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and [U], respectively). Based on those results, we selected 2h having the 1,3,5ꢀtriazine ring for
further optimization.
The previous work in our laboratory has demonstrated that in the rightꢀhand side (RHS) part,
the benzoylmorpholine moiety can be replaced with the 4ꢀmorpholinophenyl, which would result
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in better in vitro ADME property. In fact, 4ꢀmorpholinophenyl analog 3a showed a 2ꢀfold
increase in Cacoꢀ2 cell permeability compared with the benzoylmorpholine analog 2h
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P ×10 /cm·s = 7.4 and 16.3 for 2h and 3a, respectively) while the kinase inhibitory potency
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against BTK of 3a was comparable to the that of 2h (Table 2). Thus, we initiated our efforts to
explore the SAR on the leftꢀhand side (LHS) which would interact with the H3 hydrophobic
pocket of BTK while keeping the central aminotriazine ring unchanged and the RHS constant as
4ꢀmorpholinophenyl. Realizing the metabolic lability of the terminal tꢀbutyl group in 3a as we
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previously described, we focused our efforts on exploring replacement of the tꢀbutyl group to
improve the metabolic stability. Replacement with isopropyl (3b) or cyclopropyl (3c) resulted in
a slight decrease of potency for both BTK[A] and [U], although cyclopropyl analog 3c showed
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improved metabolic stability in human liver microsomes. Introduction of dimethylamino group
(3d) led to further improvement in microsomal stability in both human and mouse while the
inhibitory potency for BTK[A] was maintained. However, 6ꢀfold decrease of inhibitory potency
for BTK[U] was observed, suggesting the H3 pocket in BTK[A] and BTK[U] have different
sensitivity to substituents at the terminal benzene. Replacement of the benzene ring with thiazole
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(3e) or thiophene (3f) resulted in a significant decrease of potency for BTK[A], while the
decrease of the potency for BTK[U] was modest. Compound 3g having
tetrahydrobenzothiophene was equipotent to compound 3a, but this substitution did not improve
the metabolic stability.
Up to this point, all compound synthesized showed poor metabolic stability, presumably due to
the lability of the amide bond linking the terminal benzene in addition to the tꢀbutyl group
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metabolism that reported previously. In an attempt to improve the metabolic lability by
modifying the terminal benzene on the LHS, we next designed various cyclic amide analogs
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Table 3). Cyclization of 3a or 3c led to deterioration of the potency for both BTK[A] and [U]
compounds 3hꢀj), but this cyclization resulted in a significant improvement of the metabolic
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stabilities in both human and mice liver microsomes (compound 3i). Introduction of fluorine
atom and cyano group into the terminal tꢀbutyl benzene (3k) improved the potency while
maintaining the metabolic stability. Replacing of cyanopropane group in 3k with cyclopropyl
group (3l) further improved BTK inhibitory potency (70 nM for BTK[A]), but showed poor
stability in mouse liver microsomes. Incorporation of double bond into the cyclic amide (3m)
resulted in a remarkable improvement of BTK inhibition (vs 3l, 4.1ꢀfold and 5.3ꢀfold for
BTK[A] and BTK[U], respectively), but with moderate metabolic stability. Compound 3n
having dihydropyrrolopyrazine retained potency, but not sufficient to switch due to the metabolic
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instability. Thus, it was concluded that the dihydroisoquinolone core with cyclopropyl group and
fluorine atom was optimal for potency with acceptable metabolic stability as with the case of our
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previous report.
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Exploration of the SAR on the RHS was conducted to optimize the potency and
physicochemical property. Based on the SAR results on the pyrimidineꢀbased inhibitors
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described in the previous study, we focused our efforts on exploring substitution on the
pyrazole and pyrrole ring. Thus, a small set of Nꢀsubstituted pyrazole and pyrrole analogs was
synthesized to evaluate the potency and physicochemical property (Table 4). Replacement of the
morpholinobenzene with pyrazole as in compound 4a resulted in approximately 2ꢀfold
improvement of the inhibitory potency for BTK[A] with substantial metabolic stabilities and
aqueous solubility compared with 3m. The methyl substitution on the pyrazole (4b) led to a
further improvement of the potency for both BTK[A] and BTK[U] while maintaining good
metabolic stabilities and aqueous solubility. Further substitution of the pyrazole ring with bulky
aliphatic groups, such as cyclopropyl (4c) or cyclopropylethyl (4d) sustained BTK inhibitory
potency but the metabolic stabilities and aqueous solubilities were reduced. Substituted
cyanopyrrole (4eꢀ4i) exhibited similar inhibitory potencies but did not show the improvement in
metabolic stabilities and aqueous solubilities.
Compound 4b and 4e having Nꢀmethylꢀpyrazole and Nꢀmethylꢀcyanopyrrole, respectively,
were screened over a panel of 312 kinases to evaluate the kinase selectivity (Table S1). Both of
compounds demonstrated an excellent kinase selectivity with a similar profile, only two Tec
family kinases were inhibited at 0.3 ꢁM concentration (BMX, 80% and 84.3% inhibitions for 4b
and 4e, respectively; TEC, 99.5% and 99.2% inhibitions for 4b and 4e, respectively). TEC and
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BMX have been reported to be implicated in inflammation, this selectivity profile might be
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beneficial for the treatment of RA. On the other hand, ibrutinib was found to be inhibit both
BTK[A] and BTK[U] strongly (0.54 and 0.33 nM, respectively). Ibrutinib showed a moderate
selectivity profile (34 kinases inhibited more than 50% at 0.3 ꢁM), supporting our strategy to
obtain selective BTK inhibitors by increasing BTK[A]/BTK[U] ratio.
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As described previously, the pyrimidineꢀbased analogs showed hERG inhibition. Therefore,
safety profiling of those compounds was conducted against hERG channel (Table 4). All triazine
analogs tested showed moderate to no effects on hERG. Importantly, 4b, 4g and 4h showed
greatly reduced activities in the hERG assay. Methylpyrazole analog 4b showed an improved
hERG inhibition (IC = 24 µM) compared with the corresponding pyrimidineꢀbased analogs
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(IC50 = 14 µM). Methylpyrazole analog 4b met all of our in vitro assay criteria, and thus
selected for the further evaluations.
We next evaluated the cellular potency of 4b by measuring antiꢀIgMꢀstimulated
phosphorylation of BTK and PLCγ2 in Ramos cells. Upon BCR activation by antiꢀIgM
treatment, Tyr551 is phosphorylated by upper signal kinases, such as LYN and SYK, followed
by autophosphorylation of Tyr223 to activate BTK, then the activated BTK phosphorylates
PLCγ2 that induces intracellular calcium mobilization. As shown in Figure 3, compound 4b
potently inhibited those phosphorylations with an IC values of 19, 14 and 26 nM, respectively.
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Furthermore, 4b significantly inhibited antiꢀIgM induced B cell activation (CD86 expression) in
human PBMC with IC value of 13 nM.
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With the strong cellular potency and excellent kinase selectivity profile, compound 4b was
further evaluated in vivo. In pharmacokinetics (PK) studies of 4b using multiple species, 4b
exhibited good PK profiles with the oral bioavailabilities of 65 % in mice, 44 % in rats, and 43 %
in dogs (Table 5). Based on its adequate exposure in animals, 4b was advanced into several
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mouse models of inflammatory and autoimmune diseases to examine its in vivo efficacy. For
those animal models, we have used the following vehicle; DMSO:PEG#400:HPꢀβꢀCD
[30%(w/v)] (5:30:65), to solubilize 4b.
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Firstly, we examined the effects of 4b on FcεRIꢀmediated type I allergy in a passive cutaneous
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anaphylaxis (PCA) mouse model to evaluate the antiꢀinflammatory efficacy.
Oral
administration of 4b at 30 mg/kg significantly inhibited the PCA reaction, and its inhibition rate
was 69% as compared to vehicle control (Figure 4).
Next, we tested 4b in collagenꢀinduced arthritis (CIA) mouse model to investigate the efficacy
2
3
on autoimmune disease. Compound 4b was administered orally (15, 30 and 60 mg/kg, b.i.d.)
for 19 days, and inhibition of arthritis development was evaluated by measuring clinical arthritis
index (AI), which reflecting paw swelling and joint inflammation. As shown in Figure 5, a
marked reduction in severity of AI was observed in doseꢀdependent manner, suggesting
compound 4b attenuates arthritis development by inhibiting BTK in vivo.
Syntheses of pyrazine 2a and 1,3,5ꢀtriazine analogs 2bꢀ2h are shown in Scheme 1.
Commercially available 3ꢀaminoꢀ2ꢀmethylphenylboronic acid pinacol ester 5 was coupled with
4
ꢀtꢀbutylbenzoyl chloride to give boronate ester 6. (4ꢀAminoꢀphenyl)(morpholino)methanone 7
was coupled with 2,6ꢀdichloropyrazine by BuchwaldꢀHartwig reaction, and then the resulting
chloropyrazine 8 was coupled with boronate ester 6 by SuzukiꢀMiyaura crossꢀcoupling reaction
to give pyrazine 2a. 1,3,5ꢀTriazine analogs 2cꢀ2g were obtained in a manner similar to the
synthesis of 2a using the appropriate dichloroꢀ1,3,5ꢀtriazines. Demethylation of 2f afforded the
corresponding hydroxy triazine derivative 2g.
1
,2,4ꢀTriazine 15 was synthesized from commercially available methyl 2ꢀaminoꢀbenzoate 10.
Namely, coupling of 10 with 4ꢀtꢀbutylbenzoyl chloride, followed by hydrolysis of the methyl
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ester gave carboxylic acid 12. The carboxyl group in 12 was then converted to acetyl group using
methylmagnesium bromide via Weinreb amide 13. The resulting acetophenone 14 was oxidized
by selenium oxide, followed by cyclization with Sꢀmethylisothiosemicarbazide furnished 1,2,4ꢀ
2
4
25
0
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triazine 15. Finally, desulfurative crossꢀcoupling reaction of 15 with aniline 7 gave the
desired 1,2,4ꢀtriazine 2b.
The hydroxymethyl benzene analog 2h was synthesized from commercially available (2ꢀ
bromoꢀ6ꢀnitrophenyl)methanol 16. Protection of the hydroxyl group in 16 with TBS group,
followed by reduction of the nitro group of 17 afforded aniline 18, which was then subjected to
amide coupling with 4ꢀtꢀbutylbenzoyl chloride to give 19. The bromo group in 19 was converted
to a boronate ester group by treating with bis(pinacolato)diboron. The resulting boronate ester 20
was then coupled with 9e by Suzuki−Miyaura crossꢀcoupling, followed by TBAF treatment
yielded the desired compound 2h.
2
The general procedure for the preparation of 1,3,5ꢀtriazine derivatives with various R groups
listed in Table 2 and 3 is shown in Scheme 2, through SuzukiꢀMiyaura crossꢀcoupling of the key
chloroꢀtriazine intermediate 22. 2ꢀAminoꢀ4,6ꢀdichloroꢀ1,3,5ꢀtriazine was treated with 4ꢀ
morpholinoaniline to give common intermediate 22. The required boronate esters 23b-g, 24h-m
and 25 were prepared analogously according to the method reported literature (see supporting
information). Diamino chlorotriazine 22 was coupled with appropriate boronate esters 20, 23b-g,
2
4h-m and 25 by SuzukiꢀMiyaura crossꢀcoupling, followed by deprotection of the hydroxyl
group furnished the desired 1,3,5ꢀtriazine derivatives. In some cases (3h, 3j, and 3n), the
removal of the acetyl group was observed during the coupling reaction.
The fiveꢀmembered heterocyclic aminotriazine analogs 4a-4i were synthesized in 2 steps
starting from common intermediate 24m as shown in Scheme 3. The pyrazole derivatives 26a-
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6d and pyrrole derivative 26e-26i were readily prepared from 2ꢀaminoꢀ4,6ꢀdichloroꢀ1,3,5ꢀ
triazine and appropriately substituted aminopyrazoles or aminopyrroles as previously
1
8
described. The coupling conditions of 24m and chlorotriazines 26aꢀ26i were identical to the
one described above. Acetyl group was successfully removed by transesterification as described
in Scheme 2. For 4a, THP group was deprotected by treating with TsOH before
transesterification.
1
1
1
1
1
1
1
1
1
1
2
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2
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2
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6
7
8
9
0
CONCLUSION
Scaffold hopping strategy to eliminate the hERG inhibition was successfully applied to the
identification of novel aminotriazine derivatives as potent inhibitors of BTK. The detailed SAR
studies using two conformationally different recombinant human BTK enzymes, an activated
conformation of BTK and an unactivated conformation of BTK, led to the discovery of 4b with a
significant kinase selectivity profile. Compound 4b exhibited the strong inhibitory potency for
the BCR activation in Bꢀcells and showed excellent PK profiles in multiple species. In a mouse
CIA model, 4b significantly reduced paw swelling and joint inflammation in a doseꢀdependent
manner. Based on the in vitro potencies, PK profiles and in vivo efficacies, compound 4b was
advanced into preclinical development studies.
EXPERIMENTAL SECTION
General Procedures. Reagents and solvents were purchased from commercial sources and
used without further purification. All reactions involving airꢀ or moistureꢀsensitive reagents were
performed under a nitrogen atmosphere unless otherwise noted. Microwave reactions were run in
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a Biotage Initiator set to normal power at the indicated temperature and were performed in sealed
microwave reaction vessels. Thin layer chromatography (TLC) was carried out using Merck
GmbH Precoated silica gel 60 F254 plates. Silica gel chromatography refers to the use of an
automated medium pressure flash chromatography system (Teledyne ISCO or Yamazen Corp.)
using prepacked silica gel cartridges with UV detection at 254 nm. Preparative reverseꢀphase
HPLC (prepꢀHPLC) was performed on a Waters Autopurification system (dual triggered by
target mass and UV 254 nm) using Imtakt Unison USꢀC18, 5 mm, 50 mm × 20 mm I.D. column,
eluting with a binary solvent system A and B using a gradient elution (A, 10 mM formic acid aq.;
B, 10 mM formic acid in MeOH). All yields reported are isolated yield after removal of residual
solvents. The purity of a purified compound was determined using Shimazu Prominence HPLC
system by UV detection (215 nm) with collecting MS spectra (100ꢀ800 m/z scan) of the target
peak. The separation method is shown as following. Column: Imtakt Cadenza, 3 mm, 50 mm ×
2.0 mm I.D. Mobile phase: acetonitrile in water (10 mM formic acid) from 10% to 90% with
0
1
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8
9
0
1
2
3
4
5
6
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8
9
0
1
total 5 min run. Flow rate: 0.5 mL/min. H ꢀNMR spectra were recorded on Burker Ultra
Shield™ 400 Plus; chemical shifts (δ) are reported relative to a signal of TMS. Data format of
NMR spectra is as follows: chemical shift (δ ppm), multiplicity (s = singlet, br = broad signal, d
=
doublet, t = triplet, q = quartet, dd = doublet of doublets, dt = doublet of triplets, ddd = doublet
of doublet of doublets, m = multiplet or overlapping), coupling constant (Hz), and integration.
Very broad peaks for protons on hetero atoms are not always indicated. The synthesized
compounds for biological assay have over 95% purity, otherwise noted in each synthesis
experimental below.
4
-(t-Butyl)-N-[2-methyl-3-(6-{[4-(morpholine-4-carbonyl)phenyl]amino}pyrazin-2-
yl)phenyl]benzamide (2a). A mixture of 8 (53 mg, 0.17 mmol) and 6 (66 mg, 0.17 mmol) in
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DME (3 mL) was degassed with N for 5 min. To this mixture, Pd(PPh ) (19 mg, 0.017 mmol)
2
3 4
was added and degassed with N for 5 minutes, subsequently NaHCO (28 mg, 0.34 mmol) in
2
3
water (1 mL) was added and degassed with N for 2 min. The mixture was heated in the
2
1
1
1
1
1
1
1
1
1
1
2
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3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
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4
5
6
7
8
9
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7
8
9
0
microwave reactor at 110 °C for 20 min. The reaction mixture was extracted with ethyl acetate
(2×40 mL). The combined organic layers were washed by water (5 mL), and concentrated. The
residue was purified by flash column chromatography using 50% ethyl acetate in hexane as
1
eluent to afford 2a as offꢀwhite solids (65mg, 0.118mmol, 69%). H NMR (400 MHz, DMSOꢀ
d6) δ 9.94 (s, 1H), 9.88 (s, 1H), 8.28 (s, 1H), 8.10 (s, 1H), 8.00 – 7.92 (m, 2H), 7.85 – 7.76 (m,
2
H), 7.60 – 7.52 (m, 2H), 7.53 – 7.43 (m, 1H), 7.42 – 7.34 (m, 4H), 3.62 – 3.55 (m, 4H), 3.55 –
1
3
3.43 (m, 4H), 2.26 (s, 3H), 1.33 (s, 9H). C NMR (101 MHz, DMSO) δ 169.5, 165.8, 154.9,
1
51.4, 151.3, 142.6, 138.9, 137.8, 134.3, 133.7, 132.8, 132.2, 128.9, 128.2, 128.0, 128.0, 127.9,
+
1
26.3, 125.7, 117.8, 66.6, 35.1, 31.4, 15.8.; HRMS (ESI) m/z calculated for C H N O [M+H] :
3
3
36
5
3
550.2818, found: 550.2810, Purity 100%.
4
-(t-Butyl)-N-[2-methyl-3-(3-{[4-(morpholine-4-carbonyl)phenyl]amino}-1,2,4-triazin-5-
yl)phenyl]benzamide (2b). To a suspension of 15 (40 mg, 0.10 mmol) in toluene (2 mL) was
added 7 (25 mg, 0.12 mmol), Copper(I) 3ꢀmethylsalicylate (44 mg, 0.20 mmol), Cs CO (73 mg,
2
3
0
.22 mmol), PdOAc (2.29 mg, 10.19 nmol) and Xantphos (12 mg, 0.02 mmol). The mixture was
2
heated in the microwave reactor at 170 °C for 3 hr. The reaction mixture was filtered through a
bed of celite to remove insoluble materials. After removal of the solvent, the residue was purified
1
by prepꢀHPLC to afford 2b as offꢀwhite solids (8 mg, 0.013 mmol, 13 %). H NMR (400 MHz,
DMSOꢀd ) δ 10.47 (s, 1H), 10.03 (s, 1H), 9.09 (s, 1H), 7.96 (d, J = 8.02 Hz, 2H), 7.89 (d, J =
6
1
3
8
.31 Hz, 2H), 7.64 – 7.36 (m, 7H), 3.61 (br, 4H), 3.51 (br, 4H), 2.33 (s, 3H), 1.33 (s, 9H). C
NMR (101 MHz, DMSO) δ 169.5, 165.9, 155.4, 155.0, 150.8, 143.0, 141.4, 136.0, 134.6, 133.4,
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1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
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1
32.0, 130.7, 129.6, 129.2, 128.7, 128.0, 126.7, 125.7, 119.1, 66.6, 35.2, 31.4, 15.8.; HRMS
+
(ESI) m/z calculated for C H N O [M+H] : 551.2771, found: 551.2759, Purity 97%.
32
35
6
3
4-(t-Butyl)-N-[2-methyl-3-(4-{[4-(morpholine-4-carbonyl)phenyl]amino}-1,3,5-triazin-2-
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
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9
0
yl)phenyl]benzamide (2c). To a solution of 9c (50.0 mg, 0.16 mmol) and 6 (62 mg, 0.16 mmol)
in DME (3 mL), Pd(PPh ) (18 mg, 0.016 mmol) and K CO (43 mg, 0.31 mmol) in water (1
3
4
2
3
mL) were added, and the mixture was heated in the microwave reactor at 110 °C for 20 min. The
reaction mixture was extracted with ethyl acetate (2×40 mL), and the combined organic layers
were washed with water (5 mL), dried over sodium sulfate, filtered and concentrated. The crude
material was purified by flash chromatography on silica gel, eluted with hexane/ethyl acetate to
1
afford 2c as colorless solids (25 mg, 0.045 mmol, 29%). H NMR (400 MHz, DMSOꢀd ) δ 10.56
6
(s, 1H), 9.97 (s, 1H), 8.91 (s, 1H), 7.99 – 7.89 (m, 2H), 7.89 – 7.82 (m, 2H), 7.68 – 7.24 (m, 7H),
1
3
3
.66 – 3.54 (m, 4H), 3.56 – 3.40 (m, 4H), 2.39 (s, 3H), 1.33 (s, 9H). C NMR (101 MHz,
DMSO) δ 173.8, 168.8, 166.2, 165.3, 163.2, 154.4, 139.9, 137.3, 133.1, 132.1, 131.9, 131.9,
1
31.5, 131.4, 131.3, 128.7, 128.6, 127.9, 127.4, 125.5, 125.1, 119.8, 66.0, 34.6, 30.8, 15.4.;
+
HRMS (ESI) m/z calculated for C H N O [M+H] : 551.2771, found: 551.2770, Purity 97%.
3
2
35
6
3
Compounds 2d, 2e and 2f were synthesized from 4ꢀ(tꢀbutyl)ꢀNꢀ[2ꢀmethylꢀ3ꢀ(4,4,5,5ꢀ
tetramethylꢀ1,3,2ꢀdioxaborolanꢀ2ꢀyl)phenyl]benzamide (6) and the appropriate chlorotriazines
9
d, 9e and 9f, respectively, using a procedure similar to the procedure for the synthesis of 2c.
4
-(t-Butyl)-N-[2-methyl-3-(4-methyl-6-{[4-(morpholine-4-carbonyl)phenyl]amino}-1,3,5-
1
triazin-2-yl)phenyl]benzamide (2d). Colorless solids; 66% yield. H NMR (400 MHz, DMSOꢀ
d6) δ 10.47 (s, 1H), 9.96 (s, 1H), 7.95 (d, J = 8.45 Hz, 2H), 7.87 (d, J = 8.25 Hz, 2H), 7.67 – 7.33
1
3
(m, 7H), 3.60 (br, 4H), 3.50 (br, 4H), 2.52 (s, 3H), 2.37 (s, 3H), 1.33 (s, 9H). C NMR (101
MHz, DMSO) δ 174.4, 169.4, 165.8, 164.1, 154.9, 140.8, 137.8, 133.8, 133.7, 132.5, 132.1,
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31.9, 130.0, 129.2, 128.5, 128.0, 126.0, 125.7, 120.1, 66.6, 35.2, 31.4, 26.0, 15.9.; HRMS (ESI)
+
m/z calculated for C H N O [M+H] : 565.2927, found: 565.2936, Purity 98.4%.
3
3
37
6
3
N-[3-(4-Amino-6-{[4-(morpholine-4-carbonyl)phenyl]amino}-1,3,5-triazin-2-yl)-2-
1
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
methylphenyl]-4-(t-butyl)benzamide (2e). Colorless solids; 30.7% yield. H NMR (400 MHz,
DMSOꢀd ) δ 9.95 (s, 1H), 9.84 (s, 1H), 8.01 – 7.89 (m, 4H), 7.59 – 7.49 (m, 3H), 7.46 – 7.25 (m,
6
13
6
1
1
H), 3.60 (br, 4H), 3.51 (br, 4H), 2.37 (s, 3H), 1.33 (s, 9H). C NMR (101 MHz, DMSO) δ
74.7, 169.6, 167.3, 165.8, 164.7, 154.9, 141.9, 139.7, 137.6, 133.2, 132.2, 128.9, 128.4, 128.4,
28.0, 127.5, 125.7, 125.6, 119.5, 66.6, 35.1, 31.4, 15.8.; HRMS (ESI) m/z calculated for
+
C H N O [M+H] : 566.2880, found: 566.2874, Purity 98.9%.
32
36
7
3
4-(t-Butyl)-N-[3-(4-methoxy-6-{[4-(morpholine-4-carbonyl)phenyl]amino}-1,3,5-triazin-2-
1
yl)-2-methylphenyl]benzamide (2f). Colorless solids; 68% yield. H NMR (400 MHz, DMSOꢀ
d6) δ 10.48 (br, 1H), 9.96 (s, 1H), 8.00 – 7.90 (m, 2H), 7.92 – 7.80 (m, 2H), 7.67 (d, J = 7.95 Hz,
1H), 7.58 – 7.53 (m, 2H), 7.50 – 7.45 (m, 1H), 7.42 (d, J = 8.26 Hz, 2H), 7.37 (t, J = 7.77 Hz,
1
3
1
H), 4.01 (s, 3H), 3.60 (br, 4H), 3.50 (br, 4H), 2.39 (s, 3H), 1.33 (s, 9H). C NMR (101 MHz,
DMSO) δ 169.4, 165.8, 154.9, 140.7, 138.4, 137.9, 134.0, 132.5, 132.1, 132.0, 131.9, 130.1,
1
29.2, 128.5, 128.2, 128.0, 126.0, 125.7, 120.2, 66.6, 55.1, 35.2, 31.4, 16.0.; HRMS (ESI) m/z
+
calculated for C H N O [M+H] : 581.2876, found: 581.2883, Purity 100%.
3
3
37
6
4
4
-(t-Butyl)-N-[3-(4-hydroxy-6-{[4-(morpholine-4-carbonyl)phenyl]amino}-1,3,5-triazin-2-
yl)-2-methylphenyl]benzamide (2g). To a solution of 2f (50 mg, 0.086 mmol) in MeOH (1 mL)
was added 4N HCl in 1,4ꢀdioxane (1 mL), and the mixture was stirred at 50 °C for 4 hr. The
reaction mixture was diluted with 1N NaOH (30 mL), and extracted with ethyl acetate (3×25
mL). The combined organic layers were washed successively with water (20 mL) and brine (30
mL), dried over sodium sulfate, and concentrated. The crude material was purified by flash
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1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
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4
4
4
4
4
4
4
4
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5
5
5
5
5
5
5
5
6
chromatography on silica gel, eluted with chloroform/MeOH to afford 2g as colorless solids (7
1
mg, 0.012 mmol, 14%). H NMR (400 MHz, DMSOꢀd ) δ 12.34 (s, 1H), 10.32 (s, 1H), 10.00 (s,
6
1H), 8.03 – 7.81 (m, 3H), 7.61 – 7.48 (m, 3H), 7.47 – 7.31 (m, 4H), 3.60 (br, 4H), 3.50 (br, 5H),
1
3
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2
1
6
5
.29 (s, 3H), 1.33 (s, 9H). C NMR (101 MHz, DMSO) δ 169.4, 166.7, 165.9, 163.8, 155.9,
55.0, 140.7, 137.8, 133.7, 132.9, 132.1, 130.3, 129.8, 128.4, 128.1, 127.0, 126.3, 125.7, 120.2,
+
6.6, 35.2, 31.4, 15.5.; HRMS (ESI) m/z calculated for C H N O [M+H] : 567.2720, found:
32 35
6
4
67.2706, Purity 100%.
N-[3-(4-Amino-6-{[4-(morpholine-4-carbonyl)phenyl]amino}-1,3,5-triazin-2-yl)-2-
hydroxymethyl)phenyl]-4-(t-butyl)benzamide (2h). To a solution of 9e (256 mg, 0.764 mmol)
in DME (11 mL) was added 20 (400 mg, 0.764 mmol), Pd(PPh ) (88 mg, 0.076 mmol), K CO
(
3
4
2
3
(211 mg, 1.53 mmol) and water (3.8 mL), and the mixture was heated in the microwave reactor
at 110 °C for 20 min. The reaction mixture was diluted with water (50 mL) and extracted with
ethyl acetate (3×25 mL). The combined organic layers were washed successively with water (50
mL) and brine (50 mL), then the organic layers were dried over sodium sulfate, and
concentrated. The crude material was purified by flash chromatography on silica gel, eluted with
hexane/ethyl acetate to afford the corresponding TBS protected intermediate. The obtained
intermediate was dissolved in THF (4.1 mL), then 1 M TBAF solution in THF (0.4 mL) was
added to the solution. The mixture was stirred for 16hr at room temperature. The solvent was
evaporated, and the residue was purified by flash chromatography on silica gel, eluted with
1
hexane/ethyl acetate to afford 2h as colorless solids (62 mg, 0.11 mmol, 52%). H NMR (400
MHz, DMSOꢀd ) δ 10.41 (s, 1H), 9.85 (s, 1H), 8.09 – 7.95 (m, 1H), 7.95 – 7.83 (m, 4H), 7.65 –
6
7
.54 (m, 2H), 7.50 (dd, J = 1.44, 7.78 Hz, 1H), 7.42 (t, J = 7.85 Hz, 1H), 7.36 (d, J = 8.63 Hz,
2H), 7.29 (br, 2H), 5.67 (t, J = 5.57 Hz, 1H), 4.83 (d, J = 5.58 Hz, 2H), 3.60 (br, 4H), 3.50 (br,
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13
4
1
H), 1.33 (s, 9H). C NMR (101 MHz, DMSO) δ 174.0, 169.5, 167.0, 165.3, 164.5, 155.2,
41.7, 139.1, 138.5, 132.3, 132.1, 129.1, 128.4, 127.7, 127.5, 126.4, 126.0, 125.3, 119.6, 66.6,
+
58.4, 35.2, 31.4.; HRMS (ESI) m/z calculated for C H N O [M+H] : 582.2829, found:
3
2
36
7
4
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
5
82.2833, Purity 100%.
Compounds 3a, 3b, 3c, 3d, 3e, 3f and 3g were synthesized from 6ꢀchloroꢀN ꢀ(4ꢀ
2
morpholinophenyl)ꢀ1,3,5ꢀtriazineꢀ2,4ꢀdiamine (22) and the appropriate boronate esters using a
procedure similar to the procedure for the synthesis of 2h.
N-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
1
(
hydroxymethyl)phenyl)-4-(t-butyl)benzamide (3a). Offꢀwhite solids; 34% yield. H NMR
(400 MHz, DMSOꢀd ) δ 10.41 (s, 1H), 9.42 (br, 1H), 7.99 (s, 1H), 7.92 (d, J = 8.12 Hz, 2H),
6
7
.71 – 7.54 (m, 4H), 7.51 (d, J = 7.43 Hz, 1H), 7.42 (t, J = 7.82 Hz, 1H), 7.16 (br, 2H), 6.89 (d, J
8.60 Hz, 2H), 5.72 (br, 1H), 4.91 – 4.74 (m, 2H), 3.86 – 3.61 (m, 4H), 3.19 – 2.90 (m, 4H),
=
1
3
1.33 (s, 9H). C NMR (101 MHz, DMSO) δ 178.1, 173.6, 167.0, 165.3, 155.2, 147.2, 139.4,
1
3
9
38.5, 132.3, 131.7, 127.5, 126.5, 126.0, 125.4, 122.0, 120.7, 115.9, 110.0, 66.6, 58.4, 49.5, 35.2,
+
1.4.; HRMS (ESI) m/z calculated for C H N O [M+H] : 554.2880, found: 554.2877, Purity
3
1
36
7
3
6%.
N-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
1
(
hydroxymethyl)phenyl)-4-isopropylbenzamide (3b). Colorless solids; 15% yield. H NMR
(
400 MHz, DMSOꢀd ) δ 10.40 (s, 1H), 9.40 (br, 1H), 8.04 – 7.92 (m, 1H), 7.90 (d, J = 7.92 Hz,
6
2H), 7.71 – 7.54 (m, 2H), 7.52 – 7.34 (m, 4H), 7.17 (br, 2H), 6.88 (d, J = 8.53 Hz, 2H), 5.73 (br,
1
H), 4.91 – 4.67 (m, 2H), 3.85 – 3.59 (m, 4H), 3.12 – 2.87 (m, 5H), 1.25 (d, J = 6.94 Hz, 6H).
1
3
C NMR (101 MHz, DMSO) δ 173.6, 167.0, 165.3, 164.4, 153.0, 147.2, 139.4, 138.4, 132.7,
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1
1
1
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1
1
1
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1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
1
32.3, 127.8, 127.6, 127.1, 126.5, 125.4, 122.0, 119.9, 115.9, 66.6, 58.4, 49.5, 33.9, 24.1.;
+
HRMS (ESI) m/z calculated for C H N O [M+H] : 540.2723, found: 540.2721, Purity 100%.
3
0
34
7
3
N-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
1
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
(hydroxymethyl)phenyl)-4-cyclopropylbenzamide (3c). Offꢀwhite solids; 23% yield. H NMR
(400 MHz, DMSOꢀd ) δ 10.36 (s, 1H), 9.41 (br, 1H), 8.04 – 7.90 (m, 1H), 7.90 – 7.77 (m, 2H),
6
7
7
3
2
.60 (br, 2H), 7.49 (dd, J = 1.44, 7.80 Hz, 1H), 7.40 (t, J = 7.85 Hz, 1H), 7.28 – 7.21 (m, 2H),
.15 (br, 2H), 6.95 – 6.80 (m, 2H), 5.68 (t, J = 5.72 Hz, 1H), 4.78 (d, J = 4.94 Hz, 2H), 3.85 –
.63 (m, 4H), 3.15 – 2.92 (m, 4H), 2.11 – 1.94 (m, 1H), 1.11 – 0.96 (m, 2H), 0.82 – 0.72 (m,
1
3
H). C NMR (101 MHz, DMSO) δ 173.8, 173.6, 167.0, 165.2, 164.3, 148.8, 147.2, 139.4,
138.4, 132.2, 131.9, 127.7, 127.6, 126.5, 125.9, 125.4, 122.0, 115.9, 66.6, 58.4, 49.5, 15.7, 10.7.;
+
HRMS (ESI) m/z calculated for C H N O [M+H] : 538.2567, found: 538.2559, Purity 100%.
3
0
32
7
3
N-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
1
(hydroxymethyl)phenyl)-4-(dimethylamino)benzamide (3d). Colorless solids; 31% yield. H
NMR (400 MHz, DMSOꢀd ) δ 10.18 (s, 1H), 9.40 (br, 1H), 8.02 (d, J = 7.80 Hz, 1H), 7.88 –
6
7
.75 (m, 2H), 7.61 (br, 2H), 7.48 – 7.31 (m, 2H), 7.14 (br, 2H), 6.94 – 6.84 (m, 2H), 6.84 – 6.74
(m, 2H), 5.87 – 5.59 (m, 1H), 4.93 – 4.70 (m, 2H), 3.79 – 3.65 (m, 4H), 3.06 – 3.02 (m, 4H),
1
3
3
1
.01 (s, 6H). C NMR (101 MHz, DMSO) δ 173.7, 167.0, 165.2, 164.3, 153.0, 147.2, 139.3,
39.0, 132.3, 129.0, 127.5, 125.7, 124.8, 121.9, 121.2, 115.9, 115.1, 111.5, 66.6, 58.5, 49.5.;
+
HRMS (ESI) m/z calculated for C H N O [M+H] : 541.2676, found: 541.2668, Purity 100%.
2
9
33
8
3
N-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
(hydroxymethyl)phenyl)-5-(t-butyl)thiazole-2-carboxamide (3e). Offꢀwhite solids; 14% yield.
1
H NMR (400 MHz, DMSOꢀd ) δ 10.44 (s, 1H), 9.45 (br, 1H), 8.39 (s, 1H), 7.70 (d, J = 8.07 Hz,
6
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H), 7.66 – 7.47 (m, 3H), 7.41 (t, J = 7.85 Hz, 1H), 7.19 (br, 2H), 6.99 – 6.78 (m, 2H), 5.50 (br,
1
H), 4.70 (s, 2H), 3.81 – 3.65 (m, 4H), 3.13 – 2.94 (m, 4H), 1.42 (s, 9H).
1
3
C NMR (101 MHz, DMSO) δ 190.3, 185.3, 173.4, 166.8, 164.3, 159.5, 151.1, 147.3, 143.8,
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
39.6, 137.1, 134.4, 132.1, 127.8, 127.2, 122.1, 115.9, 66.6, 57.9, 49.5, 38.3, 30.8.; HRMS (ESI)
+
m/z calculated for C H N O S [M+H] : 561.2396, found: 561.2390, Purity 100%.
2
8
33
8
3
N-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
(hydroxymethyl)phenyl)-5-(t-butyl)thiophene-2-carboxamide (3f). Colorless solids; 44%
1
yield. H NMR (400 MHz, DMSOꢀd ) δ 10.31 (s, 1H), 9.42 (br, 1H), 7.81 (d, J = 7.91 Hz, 1H),
6
7
.68 (d, J = 3.84 Hz, 1H), 7.60 (br, 2H), 7.51 (dd, J = 1.40, 7.78 Hz, 1H), 7.39 (t, J = 7.85 Hz,
1H), 7.17 (br, 2H), 7.03 (d, J = 3.84 Hz, 1H), 6.93 – 6.84 (m, 2H), 5.63 (t, J = 5.85 Hz, 1H), 4.74
13
(d, J = 6.20 Hz, 2H), 3.80 – 3.63 (m, 4H), 3.12 – 2.94 (m, 4H), 1.38 (s, 9H). C NMR (101
MHz, DMSO) δ 173.6, 166.9, 164.3, 163.2, 160.4, 147.2, 139.5, 137.8, 136.6, 132.2, 129.0,
127.7, 126.9, 126.1, 123.5, 122.0, 115.9, 66.6, 58.2, 49.5, 40.6, 35.1, 32.4.; HRMS (ESI) m/z
+
calculated for C H N O S [M+H] : 560.2444, found: 560.2455, Purity 100%.
2
9
34
7
3
N-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
(hydroxymethyl)phenyl)-4,5,6,7-tetrahydrobenzo[b]thiophene-2-carboxamide (3g). Paleꢀ
1
yellow solids; 24% yield. H NMR (400 MHz, DMSOꢀd ) δ 10.30 (s, 1H), 9.43 (br, 1H), 7.88 (s,
6
1
H), 7.68 – 7.57 (m, 2H), 7.57 – 7.45 (m, 2H), 7.40 (t, J = 7.83 Hz, 1H), 7.17 (br, 2H), 6.89 (d, J
8.71 Hz, 2H), 5.86 (br, 1H), 4.78 (s, 2H), 3.83 – 3.63 (m, 4H), 3.18 – 2.88 (m, 4H), 2.84 – 2.69
=
13
(m, 2H), 2.69 – 2.55 (m, 2H), 1.88 – 1.69 (m, 4H). C NMR (101 MHz, DMSO) δ 167.0, 164.3,
1
1
60.4, 160.2, 147.2, 142.1, 139.4, 138.0, 136.6, 135.8, 132.2, 131.8, 130.1, 129.6, 127.6, 126.6,
22.0, 115.9, 66.6, 58.3, 49.5, 27.9, 25.3, 22.9, 22.2.; HRMS (ESI) m/z calculated for
+
C H N O S [M+H] : 558.2287, found: 558.2284, Purity 95%.
29
32
7
3
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2
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3
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5
5
5
5
6
2
-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
(
hydroxymethyl)phenyl)-5-(t-butyl)isoindolin-1-one (3h). To a solution of 22 (146 mg, 0.48
mmol) in DME (7.1 mL) was added 24h (220mg, 0.48 mmol), Pd(PPh ) (27 mg, 0.024 mmol),
3
4
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
K CO (131mg, 0.95 mmol) and water (2.3 mL), and the mixture was stirred at 100°C for 4 hr.
2
3
The reaction mixture was diluted with water (30 mL) and extracted with ethyl acetate (2×25
mL). The combined organic layers were washed successively with water (50 mL) and brine (50
mL), then the organic layers were dried over sodium sulfate, and concentrated. The crude
material was purified by flash chromatography on silica gel, eluted with hexane/ethyl acetate to
1
afford 3h as paleꢀyellow powder (35mg, 0.062 mmol, 13%). H NMR (400 MHz, DMSOꢀd ) δ
6
9.53 (br, 1H), 7.93 – 7.44 (m, 8H), 7.30 (br, 2H), 6.90 (d, J = 8.49 Hz, 2H), 5.08 (br, 1H), 4.92
13
(
s, 2H), 4.59 – 4.29 (m, 2H), 3.86 – 3.59 (m, 4H), 3.20 – 2.87 (m, 4H), 1.37 (s, 9H). C NMR
101 MHz, DMSO) δ 173.2, 167.8, 166.9, 155.7, 147.3, 143.0, 140.4, 138.9, 138.7, 132.1, 130.8,
(
130.0, 129.6, 128.5, 125.9, 123.4, 122.1, 120.6, 115.9, 98.7, 66.6, 57.5, 54.7, 49.5, 35.6, 31.6.;
+
HRMS (ESI) m/z calculated for C H N O [M+H] : 566.2880, found: 566.2867, Purity 100%.
3
2
36
7
3
2
-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
hydroxymethyl)phenyl)-6-cyclopropylphthalazin-1(2H)-one (3i). To a mixture of 24i (112
mg, 0.24 mmol) and 22 (74.6 mg, 0.24 mmol) in DME (1.6 mL) was added a solution of K CO3
(
2
(
67 mg, 0.49 mmol) in water (0.4 mL), and degassed with N for 5 min. To this mixture,
2
Pd(PPh ) (28.1 mg, 0.024 mmol) was added, and the mixture was heated in the microwave
3
4
reactor at 110 °C for 20 min. The reaction mixture was diluted with water (50 mL) and extracted
with ethyl acetate (3×25 mL). The combined organic layers were washed successively with water
(
50 mL) and brine (50 mL), then dried over sodium sulfate. The solvent was evaporated to afford
the corresponding acetylꢀprotected intermediate. The obtained intermediate was dissolved in
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MeOH/THF (2:1, 6 mL), and K CO (200 mg) were added to the solution. The mixture was
2
3
stirred at room temperature for 16 hr. The reaction mixture was diluted with water (50 mL) and
extracted with ethyl acetate (3×25 mL). The combined organic layers were washed successively
with water (50 mL) and brine (50 mL), then dried over sodium sulfate, and concentrated. The
crude residue was purified by flash chromatography on silica gel, eluted with hexane/ethyl
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
acetate to afford 3i as a paleꢀyellow powder (26 mg, 0.044 mmol, 18%). H NMR (400 MHz,
DMSOꢀd ) δ 9.50 (br, 1H), 8.46 (s, 1H), 8.18 (d, J = 8.26 Hz, 1H), 7.87 (dd, J = 1.58, 7.64 Hz,
6
1
H), 7.72 (d, J = 1.73 Hz, 1H), 7.69 – 7.50 (m, 4H), 7.49 (dd, J = 1.56, 7.86 Hz, 1H), 7.24 (br,
H), 6.90 (d, J = 8.80 Hz, 2H), 5.06 (br, 1H), 4.42 (s, 1H), 4.30 (s, 1H), 3.82 – 3.66 (m, 4H),
2
1
3
3.11 – 2.98 (m, 4H), 2.26 – 2.12 (m, 1H), 1.20 – 1.11 (m, 2H), 0.99 – 0.82 (m, 2H). C NMR
(101 MHz, DMSO) δ 173.2, 169.2, 159.2, 153.0, 151.5, 147.3, 141.9, 140.2, 138.7, 137.9, 130.8,
1
30.4, 130.3, 128.6, 128.2, 127.8, 126.6, 125.4, 123.2, 115.9, 66.6, 57.1, 49.5, 40.4, 16.0, 11.4.;
+
HRMS (ESI) m/z calculated for C H N O [M+H] : 563.2519, found: 563.2516, Purity 97%.
3
1
31
8
3
2
Compounds 3j and 3n were synthesized from 6ꢀchloroꢀN ꢀ(4ꢀmorpholinophenyl)ꢀ1,3,5ꢀ
triazineꢀ2,4ꢀdiamine (22) and the appropriate boronate esters 24j and 25, respectively, using a
procedure similar to the procedure for the preparation of 3h. Compounds 3k, 3l and 3m were
synthesized from 22 and the appropriate boronate esters 24k, 24l and 24m, respectively, a
procedure similar to the procedure for the preparation of 3i.
2
-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
(hydroxymethyl)phenyl)-6-cyclopropyl-3,4-dihydroisoquinolin-1(2H)-one (3j). Offꢀwhite
1
solids; 5.6% yield. H NMR (400 MHz, DMSOꢀd ) δ 9.50 (br, 1H), 7.86 – 7.70 (m, 2H), 7.61
6
(br, 2H), 7.52 – 7.38 (m, 2H), 7.26 (br, 2H), 7.14 – 7.03 (m, 2H), 6.95 – 6.80 (m, 2H), 5.08 (br,
1H), 4.61 – 4.47 (m, 1H), 4.42 – 4.27 (m, 1H), 4.00 – 3.79 (m, 2H), 3.81 – 3.67 (m, 4H), 3.28 –
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1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
3
2
.15 (m, 2H), 3.13 – 2.96 (m, 4H), 2.06 – 1.92 (m, 1H), 1.09 – 0.97 (m, 2H), 0.82 – 0.70 (m,
1
3
H). C NMR (101 MHz, DMSO) δ 177.9, 176.4, 165.8, 165.4, 164.0, 153.4, 149.2, 140.4,
139.6, 137.9, 132.1, 130.3, 128.2, 124.5, 124.3, 121.8, 115.9, 115.6, 113.8, 109.7, 66.6, 57.6,
+
0
1
2
3
4
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8
9
0
1
2
3
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6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
5
0.4, 49.5, 28.1, 15.8, 10.6.; HRMS (ESI) m/z calculated for C H N O [M+H] : 564.2723,
3
2
34
7
3
found: 564.2705, Purity 98%.
2
-[2-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
(hydroxymethyl)phenyl)-8-fluoro-1-oxo-1,2,3,4-tetrahydroisoquinolin-6-yl]-2-
1
methylpropanenitrile (3k). A paleꢀyellow powder; 74% yield. H NMR (400 MHz, DMSOꢀd )
6
δ 9.61 (br, 1H), 7.75 (dd, J = 2.32, 6.83 Hz, 1H), 7.61 (br, 2H), 7.54 – 7.44 (m, 2H), 7.43 (d, J =
1.86 Hz, 1H), 7.39 – 7.21 (m, 3H), 6.91 (d, J = 8.83 Hz, 2H), 4.60 (d, J = 12.04 Hz, 1H), 4.37 (d,
J = 12.02 Hz, 1H), 4.01 – 3.77 (m, 2H), 3.76 – 3.68 (m, 4H), 3.37 – 3.13 (m, 2H), 3.10 – 3.01
1
3
(
m, 4H), 1.74 (s, 6H). C NMR (101 MHz, DMSO) δ 163.3, 160.7, 160.3, 147.8, 147.7, 147.4,
143.7, 143.1, 138.0, 130.4, 129.6, 128.7, 124.3, 122.1, 120.9, 117.2, 117.2, 115.9, 113.2, 112.9,
10.0, 66.6, 57.6, 50.1, 49.5, 37.3, 28.9, 28.3.; HRMS (ESI) m/z calculated for C H FN O
3
1
3
3
34
8
+
[
M+H] : 609.2738, found: 609.2737, Purity 97%.
-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
2
(hydroxymethyl)phenyl)-6-cyclopropyl-8-fluoro-3,4-dihydroisoquinolin-1(2H)-one (3l). Offꢀ
1
white solids; 63% yield. H NMR (400 MHz, DMSOꢀd ) δ 9.50 (br, 1H), 7.76 (dd, J = 1.81, 7.34
6
Hz, 1H), 7.73 – 7.53 (m, 2H), 7.53 – 7.36 (m, 2H), 7.26 (br, 2H), 7.03 – 6.75 (m, 4H), 5.07 (br,
1H), 4.68 – 4.49 (m, 1H), 4.44 – 4.24 (m, 1H), 3.94 – 3.77 (m, 2H), 3.76 – 3.69 (m, 4H), 3.28 –
3
2
.08 (m, 2H), 3.08 – 2.94 (m, 4H), 2.06 – 1.92 (m, 1H), 1.11 – 0.98 (m, 2H), 0.88 – 0.75 (m,
1
3
H). C NMR (101 MHz, DMSO) δ 173.2, 166.9, 164.4, 163.5, 160.9, 151.7, 147.3, 143.4,
142.6, 140.4, 138.0, 132.1, 130.4, 129.6, 128.6, 122.0, 120.6, 115.9, 114.7, 112.4, 66.6, 57.7,
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Journal of Medicinal Chemistry
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5
6
7
8
9
+
5
0.2, 49.5, 28.9, 15.7, 11.0, 11.0.; HRMS (ESI) m/z calculated for C H FN O [M+H] :
3
2
33
7
3
5
82.2629, found: 582.2628, Purity 98%.
2-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
(hydroxymethyl)phenyl)-6-cyclopropyl-8-fluoroisoquinolin-1(2H)-one
(3m).
Offꢀwhite
1
solids; 68% yield. H NMR (400 MHz, DMSOꢀd ) δ 9.55 (br, 1H), 7.90 (dd, J = 1.43, 7.81 Hz,
6
1
H), 7.73 – 7.59 (m, 2H), 7.56 (t, J = 7.80 Hz, 1H), 7.45 (dd, J = 1.43, 7.95 Hz, 1H), 7.42 – 7.14
m, 4H), 6.99 (dd, J = 1.63, 13.17 Hz, 1H), 6.95 – 6.86 (m, 2H), 6.62 (dd, J = 2.09, 7.55 Hz, 1H),
.14 (br, 1H), 4.54 (dd, J = 4.98, 12.14 Hz, 1H), 4.13 (dd, J = 8.88, 12.12 Hz, 1H), 3.81 – 3.66
(
5
13
(
m, 4H), 3.12 – 2.98 (m, 4H), 2.14 – 2.00 (m, 1H), 1.14 – 1.05 (m, 2H), 0.92 – 0.81 (m, 2H). C
NMR (101 MHz, DMSO) δ 173.0, 163.7, 161.1, 158.8, 152.3, 147.3, 141.1, 140.5, 140.4, 137.8,
1
35.4, 132.1, 130.8, 130.7, 128.7, 122.1, 118.9, 115.9, 112.4, 111.4, 111.2, 104.7, 66.6, 57.2,
+
4
9.5, 15.9, 11.2.; HRMS (ESI) m/z calculated for C H FN O [M+H] : 580.2472, found:
3
2
31
7
3
580.2482, Purity 97%.
2
-(3-{4-Amino-6-[(4-morpholinophenyl)amino]-1,3,5-triazin-2-yl}-2-
(hydroxymethyl)phenyl)-7,7-dimethyl-2,3,4,6,7,8-hexahydro-1H-
1
cyclopenta[4,5]pyrrolo[1,2-a]pyrazin-1-one (3n). Offꢀwhite solids; 11% yield. H NMR (400
MHz, DMSOꢀd ) δ 9.48 (br, 1H), 7.74 (dd, J = 1.78, 7.38 Hz, 1H), 7.69 – 7.50 (m, 2H), 7.50 –
6
7
.38 (m, 2H), 7.25 (br, 2H), 6.94 – 6.83 (m, 2H), 6.57 – 6.43 (m, 1H), 5.04 (br, 1H), 4.54 (d, J =
1
1.55 Hz, 1H), 4.42 – 4.31 (m, 1H), 4.25 – 4.14 (m, 2H), 4.09 – 3.97 (m, 1H), 3.97 – 3.85 (m,
1H), 3.80 – 3.66 (m, 4H), 3.11 – 2.98 (m, 4H), 2.56 (s, 2H), 2.42 (s, 2H), 1.24 – 1.19 (m, 6H).
13
C NMR (101 MHz, DMSO) δ 159.1, 147.3, 142.9, 140.5, 140.3, 138.3, 132.1, 132.0, 131.9,
1
30.5, 129.5, 129.2, 128.4, 126.9, 125.5, 122.1, 115.9, 108.5, 66.6, 57.5, 50.3, 49.5, 45.8, 42.4,
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Journal of Medicinal Chemistry
Page 24 of 64
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9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
+
4
5
1.3, 39.5, 30.6, 30.6.; HRMS (ESI) m/z calculated for C H N O [M+H] : 581.2989, found:
32 37 8 3
81.2993, Purity 100%.
2-(3-{4-[(1H-Pyrazol-4-yl)amino]-6-amino-1,3,5-triazin-2-yl}-2-(hydroxymethyl)phenyl)-
-cyclopropyl-8-fluoroisoquinolin-1(2H)-one (4a). To a mixture of chlorotriazine 26a (124
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
6
mg, 0.42 mmol) and boronate ester 24m (200 mg, 0.419 mmol) in DME (6 mL) was added a
solution of K CO (116 mg, 0.84 mmol) in water (2 mL), and the mixture was degassed with N
2
2
3
for 5 min. Pd(PPh ) (24.21 mg, 0.021 mmol) was added to the mixture, and the mixture was
3
4
heated in the microwave reactor at 110 °C for 10 min. The reaction mixture diluted with added
water (50 mL) and extracted with ethyl acetate (3×25 mL). The combined organic layers were
washed successively with water (50 mL) and brine (50 mL), then dried over sodium sulfate. The
solvent was evaporated, and the residue was purified by flash chromatography on silica gel,
eluted with hexane/ethyl acetate to afford the corresponding acetylated product as a crude
mixture. To a solution of the obtained crude product in MeOH (10 mL), pꢀtoluenesulfonic acid
monohydrate (179 mg, 0.94 mmol) was added, and the mixture was stirred at 60°C for 1 hr.
After cooling to room temperature, the resulting solids were collected by filtration, and washed
successively with water and diethyl ether to give colorless solids. The obtained solids were
dissolved in MeOH (7 mL), and K CO (100 mg) was added to the solution. The mixture was
2
3
stirred at room temperature for 3 hr. The resulting solids were collected by filtration, and washed
successively with water and diethyl ether to afford 4a as colorless solids (150 mg, 0.30 mmol,
1
81%). H NMR (400 MHz, DMSOꢀd ) δ 12.55 (s, 1H), 9.71 (s, 1H), 8.16 – 7.59 (m, 3H), 7.61 –
6
7
.51 (m, 1H), 7.45 (dd, J = 1.45, 7.82 Hz, 1H), 7.38 – 7.31 (m, 3H), 7.29 – 7.22 (m, 1H), 7.00
(
dd, J = 1.72, 13.32 Hz, 1H), 6.67 – 6.56 (m, 1H), 5.33 – 4.93 (m, 1H), 4.60 – 4.40 (m, 1H), 4.18
13
– 4.01 (m, 1H), 2.15 – 2.00 (m, 1H), 1.17 – 1.03 (m, 2H), 0.93 – 0.81 (m, 2H). C NMR (101
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Journal of Medicinal Chemistry
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7
8
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MHz, DMSO) δ 172.7, 167.2, 163.7, 163.3, 161.1, 158.8, 158.7, 152.3, 141.1, 140.5, 140.4,
1
37.8, 135.4, 130.8, 128.6, 122.1, 118.9, 112.4, 111.4, 111.2, 104.6, 57.2, 15.9, 11.2.; HRMS
+
(ESI) m/z calculated for C H FN O [M+H] : 485.1850, found: 485.1842, Purity 97%.
25
22
8
2
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2
-(3-{4-Amino-6-[(1-methyl-1H-pyrazol-4-yl)amino]-1,3,5-triazin-2-yl}-2-
(hydroxymethyl)phenyl)-6-cyclopropyl-8-fluoroisoquinolin-1(2H)-one (4b). To a mixture of
boronate ester 24m (141 mg, 0.30 mmol) and chlorotriazine 26b (67 mg, 0.30 mmol) in DME (5
mL) was added a solution of K CO (82 mg, 0.59 mmol) in water (1.6 mL), and the mixture was
2
3
degassed with N for 5 min. Pd(PPh ) (17 mg, 0.015 mmol) was added to the mixture, and the
2
3 4
mixture was heated in the microwave reactor at 110 °C for 20 min. The reaction mixture was
diluted with water (50 mL) and extracted with ethyl acetate (3×25 mL). The combined organic
layers were washed successively with water (50 mL) and brine (50 mL), then dried over sodium
sulfate, and concentrated. The residue was purified by flash chromatography on silica gel, eluted
with hexane/ethyl acetate to afford the corresponding acetylated product as a crude mixture. To a
solution of the obtained crude product in MeOH (5 mL), K CO (100mg) was added, and the
2
3
mixture was stirred at room temperature for 2 hr. The reaction mixture was diluted with water
(50 mL) and the resulting solids were collected by filtration, and washed successively with water
1
and diethyl ether to afford 4b as offꢀwhite solids (85 mg, 0.17 mmol, 57%). H NMR (400 MHz,
DMSOꢀd ) δ 9.70 (s, 1H), 8.09 – 7.77 (m, 2H), 7.62 – 7.41 (m, 3H), 7.37 – 7.25 (m, 4H), 7.00
6
(dd, J = 1.68, 13.25 Hz, 1H), 6.62 (dd, J = 2.08, 7.52 Hz, 1H), 5.19 (dd, J = 5.00, 9.08 Hz, 1H),
4.49 (dd, J = 5.05, 12.12 Hz, 1H), 4.14 – 4.04 (m, 1H), 3.79 (s, 3H), 2.14 – 2.02 (m, 1H), 1.17 –
13
1
.01 (m, 2H), 0.92 – 0.83 (m, 2H). C NMR (101 MHz, DMSO) δ 172.7, 167.2, 163.7, 163.3,
61.1, 158.8, 152.3, 141.2, 140.5, 140.4, 137.8, 135.4, 130.9, 130.7, 128.6, 122.7, 121.9, 118.9,
1
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Page 26 of 64
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7
8
9
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
1
12.4, 111.3, 104.7, 57.2, 39.2, 15.9, 11.2.; HRMS (ESI) m/z calculated for C H FN O
2
2
6
24
8
+
[
M+H] : 499.2006, found: 499.2011, Purity 100%.
Compounds 4c, 4d, 4e, 4f, 4g, 4h and 4i were synthesized from 2ꢀ(6ꢀcyclopropylꢀ8ꢀfluoroꢀ1ꢀ
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
oxoisoquinolinꢀ2(1H)ꢀyl)ꢀ6ꢀ(4,4,5,5ꢀtetramethylꢀ1,3,2ꢀdioxaborolanꢀ2ꢀyl)benzyl acetate (24m)
and the appropriate chlorotriazines 26c, 26d, 26e, 26f, 26g, 26h and 26i, respectively, using a
procedure similar to the procedure for the preparation of 4b.
2
-(3-{4-Amino-6-[(1-cyclopropyl-1H-pyrazol-4-yl)amino]-1,3,5-triazin-2-yl}-2-
(hydroxymethyl)phenyl)-6-cyclopropyl-8-fluoroisoquinolin-1(2H)-one (4c). Colorless solids;
1
7
5% yield. H NMR (400 MHz, DMSOꢀd ) δ 9.73 (s, 1H), 8.11 (s, 1H), 7.99 – 7.83 (m, 1H),
6
7.70 – 7.50 (m, 2H), 7.50 – 7.42 (m, 1H), 7.38 (br, 2H), 7.34 (d, J = 7.34 Hz, 1H), 7.31 – 7.21
m, 1H), 7.00 (dd, J = 1.65, 13.24 Hz, 1H), 6.62 (dd, J = 2.06, 7.53 Hz, 1H), 5.22 (dd, J = 5.09,
.10 Hz, 1H), 4.51 (dd, J = 5.03, 12.10 Hz, 1H), 4.11 (dd, J = 9.16, 12.21 Hz, 1H), 3.75 – 3.59
(
9
13
(m, 1H), 2.16 – 2.01 (m, 1H), 1.19 – 0.69 (m, 8H). C NMR (101 MHz, DMSO) δ 172.7, 167.1,
1
1
63.7, 163.2, 161.1, 158.8, 152.3, 141.1, 140.5, 140.3, 137.8, 135.4, 130.8, 130.8, 128.6, 122.4,
21.0, 118.9, 112.4, 111.3, 104.7, 57.2, 33.2, 15.9, 11.2, 6.7.; HRMS (ESI) m/z calculated for
+
C H FN O [M+H] : 525.2163, found: 525.2160, Purity 99%.
28 26
8
2
2
-[3-(4-Amino-6-{[1-(1-cyclopropylethyl)-1H-pyrazol-4-yl]amino}-1,3,5-triazin-2-yl)-2-
(hydroxymethyl)phenyl]-6-cyclopropyl-8-fluoroisoquinolin-1(2H)-one (4d). Colorless solids;
1
8
3% yield. H NMR (400 MHz, DMSOꢀd ) δ 9.74 (s, 1H), 8.13 (s, 1H), 8.02 – 7.95 (m, 1H),
6
7.91 (dd, J = 1.45, 7.88 Hz, 1H), 7.67 – 7.50 (m, 2H), 7.52 – 7.42 (m, 1H), 7.42 – 7.30 (m, 2H),
7
.29 – 7.22 (m, 1H), 7.00 (dd, J = 1.64, 13.24 Hz, 1H), 6.70 – 6.56 (m, 1H), 5.18 (br, 1H), 4.69 –
4
.41 (m, 1H), 4.30 – 3.97 (m, 1H), 3.69 – 3.51 (m, 1H), 2.13 – 2.01 (m, 1H), 1.57 – 1.42 (m,
3H), 1.31 – 1.14 (m, 1H), 1.14 – 1.04 (m, 2H), 0.92 – 0.83 (m, 2H), 0.63 – 0.51 (m, 1H), 0.50 –
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Journal of Medicinal Chemistry
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7
8
9
13
0
.38 (m, 1H), 0.39 – 0.26 (m, 2H). C NMR (101 MHz, DMSO) δ 172.7, 168.7, 167.1, 163.3,
1
61.1, 158.8, 153.8, 152.2, 141.1, 140.5, 137.8, 135.4, 130.7, 130.5, 128.6, 122.1, 119.3, 118.9,
112.4, 112.4, 104.7, 62.4, 57.3, 21.0, 17.9, 15.9, 11.2, 4.3, 4.0.; HRMS (ESI) m/z calculated for
+
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
C H FN O [M+H] : 553.2476, found: 553.2452, Purity 100%.
30
30
8
2
4
-({4-Amino-6-[3-(6-cyclopropyl-8-fluoro-1-oxoisoquinolin-2(1H)-yl)-2-
(
hydroxymethyl)phenyl]-1,3,5-triazin-2-yl}amino)-1-methyl-1H-pyrrole-2-carbonitrile (4e).
1
Offꢀwhite solids; 37% yield. H NMR (400 MHz, DMSOꢀd ) δ 9.81 (s, 1H), 7.99 – 7.79 (m, 1H),
6
7
.65 – 7.49 (m, 2H), 7.49 – 7.19 (m, 5H), 7.13 – 6.83 (m, 2H), 6.62 (dd, J = 2.09, 7.53 Hz, 1H),
5
.25 – 4.95 (m, 1H), 4.62 – 4.40 (m, 1H), 4.22 – 3.93 (m, 1H), 3.75 (s, 3H), 2.15 – 2.00 (m, 1H),
13
1.16 – 1.04 (m, 2H), 0.93 – 0.81 (m, 2H). C NMR (101 MHz, DMSO) δ 172.8, 167.1, 163.5,
1
1
61.1, 158.8, 152.3, 141.2, 140.5, 140.3, 137.8, 135.4, 130.8, 128.6, 124.6, 119.8, 118.9, 114.4,
12.4, 111.4, 111.4, 111.2, 104.7, 101.1, 57.2, 35.7, 15.9, 11.2.; HRMS (ESI) m/z calculated for
+
C H FN O [M+H] : 523.2006, found: 523.2003, Purity 100%.
28
24
8
2
4
-({4-Amino-6-[3-(6-cyclopropyl-8-fluoro-1-oxoisoquinolin-2(1H)-yl)-2-
(
hydroxymethyl)phenyl]-1,3,5-triazin-2-yl}amino)-1-ethyl-1H-pyrrole-2-carbonitrile (4f).
1
Offꢀwhite solids; 79% yield. H NMR (400 MHz, DMSOꢀd ) δ 9.81 (s, 1H), 7.88 (d, J = 7.81
6
Hz, 1H), 7.67 – 7.49 (m, 2H), 7.51 – 7.41 (m, 1H), 7.42 – 7.30 (m, 3H), 7.30 – 7.22 (m, 1H),
7
.08 – 6.95 (m, 2H), 6.62 (dd, J = 2.11, 7.48 Hz, 1H), 5.25 – 5.10 (m, 1H), 4.63 – 4.43 (m, 1H),
4
.22 – 3.97 (m, 3H), 2.15 – 1.99 (m, 1H), 1.49 – 1.28 (m, 3H), 1.16 – 1.05 (m, 2H), 0.93 – 0.81
1
3
(m, 2H). C NMR (101 MHz, DMSO) δ 172.8, 167.1, 163.5, 161.1, 158.8, 152.3, 141.2, 140.5,
1
40.3, 137.8, 135.4, 130.8, 129.2, 128.7, 124.7, 118.9, 118.1, 114.4, 112.4, 111.6, 111.3, 104.7,
+
9
9.9, 57.2, 43.9, 16.8, 15.9, 11.2.; HRMS (ESI) m/z calculated for C H FN O [M+H] :
2
9
26
8
2
537.2163, found: 537.2164, Purity 100%.
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2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
4
-({4-Amino-6-[3-(6-cyclopropyl-8-fluoro-1-oxoisoquinolin-2(1H)-yl)-2-
(
hydroxymethyl)phenyl]-1,3,5-triazin-2-yl}amino)-1-isopropyl-1H-pyrrole-2-carbonitrile
1
(4g). Offꢀwhite solids; 33% yield. H NMR (400 MHz, DMSOꢀd ) δ 9.79 (s, 1H), 7.88 (d, J =
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
7
7
4
.65 Hz, 1H), 7.67 (s, 1H), 7.62 – 7.50 (m, 1H), 7.48 – 7.43 (m, 1H), 7.40 (br, 2H), 7.33 (d, J =
.38 Hz, 1H), 7.31 – 7.24 (m, 1H), 7.05 – 6.95 (m, 2H), 6.66 – 6.57 (m, 1H), 5.17 (s, 1H), 4.66 –
.33 (m, 2H), 4.19 – 4.00 (m, 1H), 2.15 – 2.00 (m, 1H), 1.48 (d, J = 6.63 Hz, 6H), 1.14 – 1.06
1
3
(
m, 2H), 0.92 – 0.82 (m, 2H). C NMR (101 MHz, DMSO) δ 163.7, 163.3, 140.6, 140.3, 135.4,
1
9
30.8, 130.7, 128.7, 118.9, 115.2, 114.5, 113.8, 112.4, 111.5, 111.4, 106.2, 105.1, 104.7, 100.0,
9.3, 87.5, 82.4, 78.1, 51.9, 51.3, 23.4, 15.9, 11.2.; HRMS (ESI) m/z calculated for C H FN O
2
3
0
28
8
+
[M+H] : 551.2319, found: 551.2310, Purity 100%.
-({4-Amino-6-[3-(6-cyclopropyl-8-fluoro-1-oxoisoquinolin-2(1H)-yl)-2-
4
(
hydroxymethyl)phenyl]-1,3,5-triazin-2-yl}amino)-1-cyclopropyl-1H-pyrrole-2-carbonitrile
1
(4h). Offꢀwhite solids; 16% yield. H NMR (400 MHz, DMSOꢀd ) δ 9.79 (br, 1H), 7.90 (d, J =
6
7
.75 Hz, 1H), 7.64 – 7.50 (m, 2H), 7.52 – 7.37 (m, 3H), 7.34 (d, J = 7.36 Hz, 1H), 7.27 (d, J =
.66 Hz, 1H), 7.06 – 6.85 (m, 2H), 6.62 (dd, J = 2.08, 7.46 Hz, 1H), 5.20 (br, 1H), 4.66 – 4.41
1
(
m, 1H), 4.25 – 4.03 (m, 1H), 3.63 – 3.46 (m, 1H), 2.15 – 2.01 (m, 1H), 1.17 – 0.96 (m, 6H),
1
3
0
1
1
.92 – 0.83 (m, 2H). C NMR (101 MHz, DMSO) δ 172.7, 167.1, 163.7, 163.3, 161.1, 158.8,
52.3, 141.2, 140.5, 140.3, 137.9, 135.3, 130.8, 128.6, 124.3, 118.9, 118.5, 114.4, 112.4, 112.2,
11.3, 104.7, 102.1, 57.2, 30.0, 15.9, 11.2, 7.0.; HRMS (ESI) m/z calculated for C H FN O
2
3
0
26
8
+
[M+H] : 549.2163, found: 549.2139, Purity 100%.
4
-({4-Amino-6-[3-(6-cyclopropyl-8-fluoro-1-oxoisoquinolin-2(1H)-yl)-2-
(
hydroxymethyl)phenyl]-1,3,5-triazin-2-yl}amino)-1-[2-(2-methoxyethoxy)ethyl]-1H-
1
pyrrole-2-carbonitrile (4i). Offꢀwhite solids; 69% yield. H NMR (400 MHz, DMSOꢀd ) δ 9.82
6
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Journal of Medicinal Chemistry
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(
s, 1H), 8.02 – 7.83 (m, 1H), 7.67 – 7.22 (m, 7H), 7.10 (d, J = 1.90 Hz, 1H), 6.99 (dd, J = 1.64,
1
3.20 Hz, 1H), 6.62 (dd, J = 2.09, 7.57 Hz, 1H), 5.27 – 5.10 (m, 1H), 4.62 – 4.44 (m, 1H), 4.24 –
4.04 (m, 3H), 3.83 – 3.63 (m, 2H), 3.57 – 3.46 (m, 2H), 3.47 – 3.35 (m, 2H), 3.23 (s, 3H), 2.14 –
13
1
1
1
1
1
1
1
1
1
1
2
2
2
2
2
2
2
2
2
2
3
3
3
3
3
3
3
3
3
3
4
4
4
4
4
4
4
4
4
4
5
5
5
5
5
5
5
5
5
5
6
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
1
2
3
4
5
6
7
8
9
0
2
1
1
1
6
.01 (m, 1H), 1.15 – 1.04 (m, 2H), 0.92 – 0.83 (m, 2H). C NMR (101 MHz, DMSO) δ 172.8,
67.1, 163.5, 161.1, 158.8, 152.3, 141.2, 140.5, 140.3, 137.8, 135.4, 130.8, 130.7, 128.6, 124.8,
18.9, 118.7, 114.4, 112.4, 111.7, 111.3, 104.7, 101.0, 71.7, 70.1, 70.1, 58.6, 57.2, 48.9, 15.9,
+
1.2.; HRMS (ESI) m/z calculated for C H FN O requires [M+H] : 611.2531, found:
3
2
32
8
4
11.2533, Purity 96%.
-(t-Butyl)-N-[2-methyl-3-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl]benzamide
(6). Under nitrogen atmosphere, 2ꢀMethylꢀ3ꢀ(4,4,5,5ꢀtetramethylꢀ1,3,2ꢀdioxaborolanꢀ2ꢀyl)aniline
(326 mg, 1.49 mmol) was dissolved in DCM (15 mL) and cooled with an ice bath. To this
4
5
solution, pyridine (0.132 mL, 1.64 mmol) and 4ꢀtꢀbutylbenzoyl chloride (439 mg, 2.23 mmol)
were added dropwise and the mixture was stirred at room temperature for 4 h. The reaction
mixture was diluted with chloroform (75 mL), washed successively with water (2×30 mL), 1N
hydrochloric acid solution (30 mL), saturated NaHCO aq. (30 mL) and brine (30 mL), then the
3
organic layer was dried over sodium sulfate, filtered and concentrated to afford 6 (565 mg, 1.44
1
mmol, 97%). H NMR (400 MHz, DMSOꢀd ) δ 9.80 (s, 1H), 7.92 (d, J = 8.3 Hz, 2H), 7.54 (m,
6
3
H), 7.39 (d, J = 7.6 Hz, 1H), 7.21 (t, J = 7.6 Hz, 1H), 2.37 (s, 3H), 1.32 (s, 9H), 1.32 (s, 12H).
+
；
LCMS (m/z): mass was calculated for C H BNO requires 393.2, found 394.0 [M+H] .
24 32
3
{4-[(6-Chloropyrazin-2-yl)amino]phenyl}(morpholino)methanone (8). A mixture of 2,6ꢀ
dichloropyrazine (250 mg, 1.68 mmol) and (4ꢀaminophenyl)morpholinꢀ4ꢀylmethanone 7 (345
mg, 1.68 mmol) in toluene (8 mL) was degassed with N gas for 5 min. Pd dba (15 mg, 0.017
2
2
3
mmol) was added to the solution and the mixture was degassed again with N for 5 min. To this
2
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