molecules Communication
The Design and Synthesis of N-Xanthone Benzenesulfonamides as Novel Phosphoglycerate Mutase 1 (PGAM1) Inhibitors Penghui Wang, Lulu Jiang, Yang Cao, Deyong Ye * and Lu Zhou * Department of Medicinal Chemistry, School of Pharmacy, Fudan University, No. 826, Zhangheng Rd., Shanghai 201203, China;
[email protected] (P.W.);
[email protected] (L.J.);
[email protected] (Y.C.) * Correspondence:
[email protected] (D.Y.);
[email protected] (L.Z.); Tel.: +86-21-5198-0117 (D.Y.); +86-21-5198-0125 (L.Z.)
Received: 13 May 2018; Accepted: 4 June 2018; Published: 8 June 2018
Abstract: Upregulation of phosphoglycerate mutase 1 (PGAM1) has been identified as one common phenomenon in a variety of cancers. Inhibition of PGAM1 provides a new promising therapeutic strategy for cancer treatment. Herein, based on our previous work, a series of new N-xanthone benzenesulfonamides were discovered as novel PGAM1 inhibitors. The representative molecule 15h, with an IC50 of 2.1 µM, showed an enhanced PGAM1 inhibitory activity and higher enzyme inhibitory specificity compared to PGMI-004A, as well as a slightly improved antiproliferative activity. Keywords: phosphoglycerate mutase 1; inhibitors; anti-cancer; xanthone
1. Introduction Metabolic reprogramming has been considered as one of 10 essential hallmarks of cancer cells [1]. This metabolic phenotype is associated with the phenomenon of cancer cells altering their metabolic pathways, including bioenergetics and anabolic biosynthesis, to satisfy the anabolic demands of macromolecular biosynthesis and to maintain cellular redox homeostasis in response to the escalated production of toxic reactive oxygen species (ROS) during cell proliferation [2]. The first identified cancer cell metabolism reprogramming phenomenon was the Warburg effect [3], which refers to cancer cells relying on the high rate of aerobic glycolysis to produce energy rather than the efficient mitochondrial oxidative phosphorylation as in most normal cells [4]. This specific metabolic pattern in cancer cells serves to supply glycolytic intermediates as building blocks for anabolic biosynthesis of macromolecules, such as RNA/DNA, proteins, and lipids [5]. More and more researchers have focused on the key enzymes in cancer cell metabolism reprogramming to find new cancer therapeutic targets [6–10]. Phosphoglycerate mutase 1 (PGAM1) is a glycolytic enzyme that catalyzes the interconversion of 3-phosphoglycerate (3PG) and 2-phosphoglycerate (2PG) with 2,3-bisphosphoglycerate (2,3-BPG) as a cofactor in glycolysis. PGAM1 was found to be upregulated in a variety of human cancers, including breast cancer [11], prostate cancer [12], lung cancer [13], etc. A recent work by Hitosugi showed that in cancer cells, upregulated PGAM1 coordinates glycolysis and biosynthesis to promote cancer cell proliferation and tumor growth [14]. The molecular mechanism of this function of PGAM1 is that the upregulation of PGAM1 leads to a lower intracellular level of 3PG and a higher intracellular level of 2PG, which results in a high level of pentose phosphate pathway (PPP) flux and activated serine synthesis pathway (SSP), respectively [14,15]. This mechanism facilitates the conversion of glycolytic intermediates to the precursors of amino acids and ribose, which are building blocks of DNA/RNA and proteins. In addition, both downregulation of PGAM10 s expression and inhibition Molecules 2018, 23, 1396; doi:10.3390/molecules23061396
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DNA/RNA and and proteins. proteins. In In addition, addition, both both downregulation downregulation of of PGAM1′s PGAM1′s expression expression and and inhibition inhibition of of DNA/RNA of its metabolic activity has been shown to attenuate cell proliferation and tumor growth [14,16]. its metabolic activity has been shown to attenuate cell proliferation and tumor growth [14,16]. its metabolic activity has been shown to attenuate cell proliferation and tumor growth [14,16]. Accordingly, not only reduce thethe indispensable energy supply for Accordingly,developing developingPGAM1 PGAM1inhibitors inhibitorscould could not only reduce the indispensable energy supply Accordingly, developing PGAM1 inhibitors could not only reduce indispensable energy supply cancer cellscells but also the required anabolic processes including PPP andPPP SSP,and and SSP, hence provide a for cancer cancer cells but block also block block the required required anabolic processes including PPP and SSP, and and hence for but also the anabolic processes including hence new dual-functional anticancer strategy [17]. provide aa new new dual-functional dual-functional anticancer anticancer strategy strategy [17]. [17]. provide Although PGAM1 has been been identified as one potential anticancer target, only three small Although PGAM1 PGAM1 has has been identified identified as as one one potential potential anticancer anticancer target, target, only only three three small small Although molecules have currently been discovered as PGAM1 inhibitors (Figure 1). was initially been discovered as PGAM1 inhibitors (Figure 1). MJE3 molecules have currently been discovered as PGAM1 inhibitors (Figure 1). MJE3 was initially screened for for inhibiting inhibiting the the proliferation proliferation of of MDA-MB-231 MDA-MB-231 cells cells and and its its antiproliferation antiproliferation effect effect was was screened subsequently confirmed come from its inhibitory inhibitory activity activity against against PGAM1 PGAM1 through through in in situ subsequently confirmed confirmed to to come come from from its situ proteome proteome subsequently to reactivity profiling [18]. A reactivity profiling [18]. further study showed that MJE3 could inactivate PGAM1 through covalent reactivity profiling [18]. A further study showed that MJE3 could inactivate PGAM1 through covalent 0 s K100 by its spiroepoxide substructure [19]. PGMI-004A, with moderate modification of PGAM1 PGAM1′s K100 by its its spiroepoxide spiroepoxide substructure substructure [19]. [19]. PGMI-004A, with moderate moderate modification of PGAM1′s enzymatic showed a significant antiproliferation on both a cellular level and mice activity, effect enzymatic activity, showed a significant antiproliferation effect on both a cellular level and mice xenograft models [14]. Additionally, the third PGAM1 inhibitor, epigallocatechin gallate (EGCG), models [14]. [14]. Additionally, Additionally, the the third third PGAM1 PGAM1 inhibitor, inhibitor, epigallocatechin epigallocatechin gallate gallate (EGCG), (EGCG), has has xenograft models has recently been reported [20]. Although EGCG is the most potentPGAM1 PGAM1inhibitor inhibitorat atthe the molecular molecular recently been reported [20]. Although EGCG is the the most potent PGAM1 inhibitor at the recently been reported [20]. Although EGCG is most potent level, its its polyphenol polyphenol structure structure structure and and off-target off-target effect effect may may limit limitits itsfurther furtherapplications applications[21]. [21]. level, and off-target effect may limit its further applications [21]. OH OH
O O O O O O
OH OH
O O
H H N N
O O
O O
O O
O O
NH NH
O O
OH OH OH OH O O S S N O N H O H
O O
HO HO
OH OH
CF33 CF OH OH
O O
OH OH
O O
OH OH MJE3 IC IC50== 33 33 uM uM MJE3 50
PGMI-004A PGMI-004A
EGCG EGCG
OH OH
PGAM1: IC IC50 = 13.1 13.1 uM uM 50= PGAM1: 25.6 uM uM H1299 cell cell :: IC IC50== 25.6 H1299 50
PGAM1: IC IC50 = 0.49 0.49 uM uM 50= PGAM1: H1299 cell: cell: IC IC50 > 80 80 uM uM 50 > H1299
Figure 1. 1. The The inhibitors inhibitors of of PGAM1. PGAM1. Recently, Recently, starting from from the the PGMI-004A, PGMI-004A, we we conducted conducted scaffold scaffold Figure inhibitors Recently, starting starting hopping and and aa sulfonamide sulfonamide reversal reversal strategy strategy to to discover discover aa series series of of 1,2,8-trihydroxy 1,2,8-trihydroxy xanthone xanthone hopping hopping and sulfonamide reversal strategy to discover derivatives as as novel novel PGAM1 PGAM1 inhibitors inhibitors (Figure (Figure 2). 2). The The inhibitory activities both on the enzymatic and derivatives inhibitory activities both on the enzymatic The inhibitory activities the enzymatic and and cellular level of those xanthone derivatives were significantly improved compared to PGMI-004A cellular level of ofthose thosexanthone xanthonederivatives derivatives were significantly improved compared to PGMI-004A were significantly improved compared to PGMI-004A [22]. [22]. Besides, Besides, the xanthone xanthone core as an an important important scaffold withbiologic diverseactivities, biologic activities, activities, such as as [22]. the as with diverse biologic such Besides, the xanthone core ascore an important scaffold scaffold with diverse such as antitumor, antitumor, antioxidant, antioxidant, anti-inflammation, etc., was of of documented documented relevance to human human diseases [23– antitumor, anti-inflammation, was relevance to diseases antioxidant, anti-inflammation, etc., was ofetc., documented relevance to human diseases [23–27]. In[23– this 27]. In In this this paper, maintaining maintaining the xanthone xanthoneand scaffold and considering considering that the the ortho-dihydroxy ortho-dihydroxy phenol paper, maintaining the xanthone scaffold considering that the ortho-dihydroxy phenol phenol moiety 27]. paper, the scaffold and that moietycause mightmetabolic cause metabolic metabolic instability [28], we we removed removed the C2-hydroxy C2-hydroxy group and to design design and might instability [28], we removed the C2-hydroxy group to design synthesize moiety might cause instability [28], the group to and synthesize a series of new N-xanthone benzenesulfonamides. What is more, the SAR of the A-ring has a series of anew N-xanthone benzenesulfonamides. What isWhat more,is the SAR theofA-ring has also synthesize series of new N-xanthone benzenesulfonamides. more, theof SAR the A-ring has also been been explored. been explored. also explored.
Figure 2. 2. The The optimization optimization process process of of PGMI-004A. PGMI-004A. Figure Figure 2. The optimization process of PGMI-004A.
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2. Results and Discussion Molecules 2018, 23, x FOR PEER REVIEW 3 of 14 2. Results and Discussion Molecules 2018, xthe FOR PEER REVIEWeffects influencing the activities on the benzene3 ring, 3 of 14 First, to evaluate substituent Molecules 2018, 23, x23, FOR PEER REVIEW of 14 14 Nxanthone were synthesized as shown inthe Scheme 1. Theonxanthone core 3 ring, was 2. benzenesulfonamides Results and Discussion First, to evaluate the substituent effects influencing activities the benzene 2.2.Results and Discussion Results and Discussion constructed by treating 2,6-dihydroxybenzoic acid 1 and phloroglucinol 2 with Eaton′s reagent and 14 N-xanthone benzenesulfonamides were synthesized as shown Schemeon 1. The xanthonering, core First, to evaluate the substituent effects influencing theinactivities thering, benzene 143 NFirst, to evaluate the substituent effects influencing the activities on the benzene 14 N0 converted to triflate 4 by reacting with Tf 2O.effects After pivaloylation of triflate 4, the dipivalate 5 was First, to evaluate the substituent influencing the activities on the benzene ring, 14 Nwas constructed by treating 2,6-dihydroxybenzoic acid 1asand phloroglucinol withxanthone Eaton s reagent xanthone benzenesulfonamides were synthesized shown in 1. Scheme 1.2 The 3 was xanthone benzenesulfonamides were synthesized as shown in Scheme The xanthone core 3 was core xanthone benzenesulfonamides were synthesized as shown in Scheme 1. The xanthone core 3 obtained and was then converted to xanthone amine 7 through the Buchwald amination-hydrolysis and converted to triflate 4 by2,6-dihydroxybenzoic reacting with Tf2 O. After of2 with triflate the dipivalate 5 waswas constructed treating 2,6-dihydroxybenzoic 1 and phloroglucinol 24,with Eaton′s and constructed by by treating acid 1 acid andpivaloylation phloroglucinol Eaton′s reagent andreagent constructed by treating 2,6-dihydroxybenzoic acid 1 and phloroglucinol 2 with Eaton′s reagent protocol [29]. The xanthone amineto7 xanthone was treated with7diverse substituted benzenesulfonyl chloridesand obtained and was then converted through thetriflate Buchwald amination-hydrolysis converted to to triflate 4 by4 reacting with with Tf2O. amine After of the dipivalate was converted triflate by reacting Tf2O. pivaloylation After pivaloylation of4, triflate 4, the 5dipivalate 5 was converted to triflate 4 to byafford reacting with Tf 2O. After pivaloylation of triflate 4, the dipivalate 5 was and followed deprotection theto target molecules N-xanthone benzenesulfonamides 9a–9n. obtained and waswas thenthen converted xanthone amine 7 diverse through the Buchwald amination-hydrolysis protocol [29]. by The xanthone amine 7 to was treated with substituted benzenesulfonyl chlorides obtained and converted xanthone amine 7 through the Buchwald amination-hydrolysis obtained and was then converted to xanthone amine 7 through the Buchwald amination-hydrolysis protocol [29]. TheThe xanthone 7 was with diverse substituted benzenesulfonyl chlorides 9a–9n. and followed by deprotection toamine afford the7treated target molecules benzenesulfonamides protocol [29]. xanthone amine was treated with N-xanthone diverse substituted benzenesulfonyl chlorides protocol [29]. The xanthone aminethe7 target was treated with diverse benzenesulfonamides substituted benzenesulfonyl chlorides and followed by deprotection to afford molecules N-xanthone 9a–9n. and followed by deprotection to afford the target molecules N-xanthone benzenesulfonamides 9a–9n. and followed by deprotection to afford the target molecules N-xanthone benzenesulfonamides 9a–9n.
SchemeScheme 1. Reagents and and conditions: reagent(7.7 (7.7 2O5 solution in 3MeSO H), 1. Reagents conditions:(a) (a)Eaton’s Eaton’s reagent wtwt % P% 2OP 5 solution in MeSO H), 80 3°C ; 80 °C; Scheme 1. Reagents andDCM, conditions: (a) reagent (7.7 wt %THF; P2THF; O5(d) solution in 2MeSO 80 ◦ C; Tfpyridine, 2O, pyridine, DCM,0 0°C°C (c) NaH NaH (60%), PivCl, Pd(OAc) , BINAP, (b) Tf(b) 2O, ; ; Eaton’s (c) (60%), PivCl, (d) Pd(OAc) 23, H), BINAP, ◦ Scheme 1. Reagents and conditions: (a) Eaton’s reagent (7.7 wt % P 2 O 5 solution in MeSO 3 80 °C; 2CO 3NaH , dioxane, reflux; (e) THF, H2O, r.t.; r.t.; (f) BINAP, RSO 2Cl, diphenylmethanimine, pyridine, r.t.; (g) H),(g) diphenylmethanimine, Cs (b) Tf2 O, pyridine,1.DCM, 0 2CO C;and (c) (60%), PivCl, THF; (d) Pd(OAc) 3, dioxane, reflux; (e)HCl, HCl, THF, H 2O, (f) 2Cl, pyridine, diphenylmethanimine, Cs Scheme Reagents conditions: (a) Eaton’s reagent (7.7 wt2 ,% P2ORSO 5 solution in MeSOr.t.; 3H), 80 °C; NaOH, MeOH, H2O, (e) r.t. HCl, Tf 2O, pyridine, DCM, 0 O, °C; (c) NaH Cl, (60%), PivCl, THF; (d) MeOH, Pd(OAc) Cs , (b) dioxane, THF, H (f) RSO pyridine, r.t.; (g) NaOH, H222,,O,BINAP, r.t. 2 CO3 MeOH, NaOH, O,pyridine, r.t. (b) Tf2H O,2reflux; DCM, 0 2 °Cr.t.; ; (c) NaH2 (60%), PivCl, THF; (d) Pd(OAc) BINAP, diphenylmethanimine, Cs2CO3, dioxane, reflux; (e) HCl, THF, H2O, r.t.; (f) RSO2Cl, pyridine, r.t.; (g) 3, dioxane, reflux; HCl, THF, H2O, (f) RSO 2Cl,inhibitory pyridine, r.t.; (g) diphenylmethanimine, Cs2COassay A NaOH, multiple-enzymes system [14] (e) was conducted to r.t.; measure the MeOH, H2O,coupled r.t. A multiple-enzymes coupled assay system [14] was conducted to measure thewith inhibitory NaOH, MeOH, H 2 O, r.t. A multiple-enzymes coupled assay system [14] was conducted to measure the inhibitory activities activities of the abovementioned compounds against PGAM1 (Table 1). The compounds
activities of the abovementioned against PGAM1 (Table 1). The compounds with unsubstituted benzene (9a and 9b)compounds or benzene monosubstituted with halogens of low atomic weight of the abovementioned compounds against PGAM1 (Table The compounds with unsubstituted A multiple-enzymes coupled assay system [14]1). was conducted to measure the inhibitory A multiple-enzymes coupled assay system [14] was conducted to measure the inhibitory (9c, 9d, and 9e) showed very low inhibitory activities against PGAM1 (inhibitory ratios at 20 uM were unsubstituted benzene (9a and 9b) or benzene monosubstituted with of low atomic weight benzene (9a and 9b) monosubstituted with halogens of lowhalogens atomic 9d, and 9e) activities of or thebenzene abovementioned compounds against PGAM1 (Table weight 1). The(9c, compounds with less than 50%), suggesting that the removal of the C2-hydroxy group would reduce the inhibitory activities of the abovementioned compounds against PGAM1 (Table 1). The compounds with (9c, 9d, and 9e) showed very low inhibitory activities against PGAM1 (inhibitory ratios at 20 uM were showed unsubstituted very low inhibitory activities against PGAM1monosubstituted (inhibitory ratios at 20 uM were less atomic than 50%), benzene (9a and 9b) or benzene with halogens of low weight activity. However,benzene the iodo-substituted compound (9f) and the di-halogenated benzene compounds unsubstituted (9a and 9b) or benzene monosubstituted with halogens of low atomic weight less than(9c, 50%), suggesting that theC2-hydroxy removal ofgroup the C2-hydroxy group reduce the at inhibitory suggesting that the removal of very the would reduce the would inhibitory activity. However, 9d,and and showed low inhibitory activities against PGAM1 (inhibitory ratios 20 uM were (9g, 9h, 9i)9e) restored the inhibitory activities, which indicating that bulky groups with enhanced (9c, 9d, and 9e) showed very low inhibitory activities against PGAM1 (inhibitory ratios at 20 uM were activity. However, the iodo-substituted compound (9f) and the di-halogenated benzene compounds the iodo-substituted compound (9f) and the di-halogenated benzene compounds (9g, 9h, 9i) less than 50%), the removalactivity of theand C2-hydroxy group would reducedue theand inhibitory lipophilicity might suggesting be beneficial that for the inhibitory counteract the decrease in activity less than 50%), suggesting that the removal of theindicating C2-hydroxy group would reduce the inhibitory (9g, 9h, and 9i) restored the inhibitory activities, which that bulky groups with enhanced theinhibitory removal ofactivities, thethe C2-hydroxyl group. Thus, large groups including (tert-butyl)phenyl (9k),compounds However, iodo-substituted compound (9f)groups and the di-halogenated benzene restoredtoactivity. the which indicating that bulky with enhanced lipophilicity might activity. However, the iodo-substituted compound (9f) and the di-halogenated benzene compounds cyclohexylphenyl (9l), biphenyl and naphthalenyl (9n)and were introduced to the benzene ring of enhanced lipophilicity might beneficial for(9m), the and inhibitory activity counteract the decrease in activity due (9g, 9h, 9i) restored the inhibitory activities, which indicating that bulky groups with be beneficial forand thebe inhibitory activity counteract the decrease in activity due to the removal of (9g,benzenesulfonamides. 9h, and 9i) restoredAs theexpected, inhibitory activities, which indicating that bulky groups with enhanced the the inhibitory activities of the compounds with large to the removal of the C2-hydroxyl group. Thus, large groups including (tert-butyl)phenyl (9k), lipophilicity might be beneficial for the inhibitory activity and counteract the decrease in activity the C2-hydroxyl group. Thus, large groups including (tert-butyl)phenyl (9k), the cyclohexylphenyl (9l),due lipophilicity might be beneficial for the inhibitory activity and counteract decrease in activity due substituents is significantly increased compared to methyl substituted compounds 9j. Among them, cyclohexylphenyl (9l), biphenyl (9m), and naphthalenyl (9n) were introduced to the benzene ring of to the removal of the C2-hydroxyl group. Thus, large groups including (tert-butyl)phenyl (9k), biphenylto(9m), and naphthalenyl (9n) were introduced to the benzene ring of the benzenesulfonamides. the of the C2-hydroxyl group. Thus, groups including (tert-butyl)phenyl (9k), 9m (IC 50 =removal 5.5 ± 1.1 μM), containing a biphenyl moiety, was large the most effective PGAM1 inhibitor with cyclohexylphenyl (9l), (9m), andinhibitory naphthalenyl (9n) were to the benzene ring of theexpected, benzenesulfonamides. Asbiphenyl expected, the of introduced the compounds with large As inhibitory activities the compounds with activities large substituents significantly increased value 2-fold lower than theofreference inhibitor PGMI-004A. In were general, theis substituent effects an IC50the cyclohexylphenyl (9l), biphenyl (9m), and naphthalenyl (9n) introduced to the benzene ring of the Aswith expected, themethyl inhibitory activities of the compounds with large substituents is significantly increased compared to substituted 9j. Among them, compared tobenzenesulfonamides. methyl substituted compounds 9j. Among them, 9m (IC50 compounds = 5.5 ± 1.1 µM), containing a ofthe the benzene ring was consistent our previous benzenesulfonamides. As expected, the work. inhibitory activities of the compounds with large substituents is significantly increased compared to methyl substituted compounds 9j. Among them, 9m (IC50substituents =moiety, 5.5 ± 1.1was μM), containing a biphenyl moiety, was the most effective PGAM1 inhibitor with biphenyl the most effective PGAM1 inhibitor with an IC value 2-fold lower than the 50 is significantly increased compared to methyl substituted compounds 9j. Among them, 9m (IC 50 = 5.5 ± 1.1 μM), containing a biphenyl wasPGAM1. theInmost effective PGAM1 inhibitor with Table Inhibitory activities of moiety, 9a–9n against 2-fold lower than the reference inhibitor PGMI-004A. general, thering substituent effects an IC50 value reference inhibitor PGMI-004A. In1.general, substituent effects of most the benzene was consistent 9m (IC 50 = 5.5 ± 1.1 μM), containing a the biphenyl moiety, was the effective PGAM1 inhibitor with 50 value 2-fold lower than the reference inhibitor PGMI-004A. In general, the substituent effects an IC of theour benzene was consistent with previous work. PGMI-004A. In general, the substituent effects with previous work. value 2-fold lower than theour reference inhibitor an IC50 ring of the benzene ring was consistent with our previous work. of the benzene ring was consistent with our previous work. Table 1. Inhibitory Inhibitory activities activities of of 9a–9n 9a–9n against against PGAM1. PGAM1. Table 1. Table 1. Inhibitory activities of 9a–9n against PGAM1. * Entry Table Compounds R of 9a–9n against IC50 (μM) 1. Inhibitory activities PGAM1. 1
9a
>20 **
2
9b
>20
Entry 3 Compounds Entry 9c Entry Compounds Compounds Entry Compounds 9a 1 1 1 9a 9a 9a 1 2 2
3
2 2
9b
3 3
9c
9b
9b 9b 9c 9c
R
R R R
* (µM) * IC50 (μM) IC(μM) >20 50 * IC50 IC50 (μM) * >20 **>20 ** >20 ** >20 ** >20 >20 >20>20 >20 >20 >20
Table 1. Inhibitory activities of 9a–9n against PGAM1.
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Compounds
1
9a
2
Table 1. Cont. 9b
>20
9c
>20>20
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IC50 (μM) *
Entry
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>20 **
4 of 14 4 of 14 4 of 14 4 of 14 4 of 14 4 of 14 4 of 14 4 of 14 4 of 14 4 of 14 4 of 14
9d >20>20 9d >20 9d >20 9d >20 9d >20 9e >20 >20 9e >20 54 5 9e 9d >20 9d >20 9e >20 9d 455 >20 9e 9f 645 10.1>20 ± 0.5 9d 9e 9f 10.1 ± 0.5 9d 9e >20 564 9f 10.1 ± 0.5 6 9f 9d 10.1 ± 0.5 465 >20 9e 9f 10.1 ± 0.5 9e >20 566 9f 10.1 ± 0.5 9e 9f 10.1>20 0.5 9g 7655 13.2 ±± 0.4 9e >20 9f 10.1 0.5 9g 76 13.2 ±± 0.4 9e >20 9f 657 10.1 ± 0.5 9g 13.2 0.4 9f 10.1 0.5 9g 76 13.2 ± 0.4 7 9g 13.2 ± 0.4 9f 67 10.1 9g 13.2 ± 0.5 0.4 9f 67 10.1 9g 13.2 ± 0.5 0.4 9g 13.2 0.4 9h 87 6.4 ±±0.4 9g 78 13.2 0.4 9h 6.4 ±± 0.4 9g 13.2 0.4 9h 87 6.4 ±±0.4 9g 13.2 0.4 9h 87 6.4 ±±0.4 78 13.2 0.4 9h 6.46.4 ±± 0.4 8 9h 9g ± 0.4 9h 8 6.4 ± 0.4 9h 6.4 ±± 0.4 9i 10.2 0.7 98 9h 89 6.4 ±±0.4 9i 10.2 0.7 9h 6.4 ±±0.4 9i 10.2 0.7 98 9h 6.4 ±±0.4 9i 10.2 0.7 98 9h 89 6.4 ±±0.4 9i 10.2 0.7 9i 10.2 ± 0.7 9 9 9i 10.2 ± 0.7 9j 10 >20 9i 10.2 ± 0.7 9 9j 10 >20 9i 10.2 ± 0.7 9 9j 10 >20 9i 10.2 0.7 9 9j 10 9k 8.4>20 ±±±2.7 11 9i 10.2 0.7 9 9j 10 >20 9k 8.4 ±±2.7 11 9i 10.2 0.7 9 9j 10 >20 9k 8.4 ± 2.7 11 9j 10 >20 10 9j 9k >20 8.4>20 2.7 11 9l 12 5.9 ±±± 1.2 9j 10 9k 8.4>20 2.7 11 9l 12 5.9 ± 2.7 1.2 9j 10 9k 8.4 ± 11 9l 12 5.9 ±± 1.2 9j 10 >20 9k 8.4 2.7 11 9l 12 5.9 ± 1.2 9j 10 >20 5.5 1.1 13 9k 8.4 ±± 2.7 11 11 9k 9m ± 2.7 9l 12 5.98.4 1.2 9m 5.5 1.1 13 9k 8.4 2.7 11 9l 12 5.9 ±±± 1.2 9k 8.4 2.7 11 9m 5.5 1.1 13 9l 12 5.9 ±± 2.7 1.2 9m 5.5 1.1 13 9k 8.4 11 9l 12 5.9 ±± 1.2 9m 5.55.9 1.1 13 9l 12 5.9 1.2 12 9l 9m ± 1.2 9n 6.0 ± 0.6 14 5.5 1.1 13 9l 12 5.9 ±± 0.6 1.2 9n 6.0 14 9m 5.5 1.1 13 9l 12 5.9 1.2 9n 6.0 ±± 1.1 0.6 14 9m 5.5 13 9n 6.0 ±± 0.6 14 9m 5.5 1.1 13 9n PGMI-004A 6.0 0.6 14 15 135.5 ±± 0.1 5.5 1.1 13 13 9m 9m ± 1.1 9n 6.0 ± 0.6 14 15 13 ± 0.1 9m PGMI-004A 5.5 1.1 13 9n± SD 0.6 14 15 PGMI-004A 13The ±± 0.1 * All IC50 values are reported as means from three determinations. **6.0 inhibition ratios at 20 15 PGMI-004A 13The ± 0.1 9n± SD 6.0 0.6 14 * All IC50 values are reported as means from three determinations. ** inhibition ratios at 20 15 PGMI-004A 13The ±± 0.1 9n± SD 6.0 0.6 14 * Allwere IC50 values are50%. reported as means from three determinations. ** inhibition ratios at 20 μM less than 9n± SD 15 PGMI-004A 13The ±± 0.1 0.6 14 * Allwere IC50 values are50%. reported as means from three determinations. **6.0 inhibition ratios at 20 μM less than 9n 15 PGMI-004A 13 ± 0.1 9n 6.0 ± 0.6 6.0 0.6 14 *μM Allwere IC5014 values are reported as means ± SD from three determinations. ** The inhibition ratios at 20 less than 50%. 15 PGMI-004A ± 0.1 *μM Allwere IC50 values are50%. reported as means ± SD from three determinations. **13The inhibition ratios at 20 less than 15 of 9maswas PGMI-004A ± 0.1 *μM Allwere IC50modification values are50%. reported means ± SD from three determinations. **13The inhibition ratios at 20 that less than Further conducted at the A-ring of the xanthone core. Considering 15 PGMI-004A ± 0.1 *μM Allwere IC50 modification values are50%. reported means ± SD from determinations. **13The inhibition ratios at 20 that less than Further 9maswas conducted atthree the A-ring of the xanthone core. Considering 15 of PGMI-004A ± 0.1 *μM Allwere IC5015 values are50%. reported aswas means ± SD from three determinations. **13The inhibition ratios lessof than Further modification of 9mderived conducted at the A-ring of the xanthone core. Considering that the substituents A-ring was from the starting material substituted salicylic acids,at a20series PGMI-004A 13 0.1 * Allwere IC50modification values are50%. reported aswas means ± SD the from three determinations. ** The± inhibition ratios ata20series Further of 9mderived conducted at the A-ring of the xanthone core. Considering that μM less than the substituents of A-ring was from starting material substituted salicylic acids, * All IC 50 values are reported as means ± SD from three determinations. ** The inhibition ratios at 20series Further modification of 9m was conducted at the A-ring of the xanthone core. Considering that μM were less than 50%. the substituents of A-ring was derived from the starting material substituted salicylic acids, a of A-ring substituted compounds were synthesized according to Schemes 2 and 3 and their enzymatic * All IC values are reported as means ±derived SD from three determinations. ** The ratios µM were less Further modification of 9m was conducted the A-ring ofinhibition the xanthone core. Considering that μM were less 50%. 50 the substituents ofthan A-ring was from the at starting material substituted acids, a series of A-ring substituted compounds were synthesized according to Schemes 2 andatsalicylic 320and their enzymatic Further modification of 9m was conducted at the A-ring of the xanthone core. Considering that μM were less than 50%. the substituents of A-ring was derived from the starting material substituted acids, a series of A-ring substituted compounds were synthesized according to Schemes 2(Table andsalicylic 3 and their enzymatic than 50%. inhibitory activities were evaluated by the previous enzyme assay method 2). Further analysis
Further modification of 9mderived was conducted the A-ring of Schemes the xanthone core. that the substituents of A-ring was from the at starting material substituted acids, a series of A-ring substituted compounds were synthesized according to 2 (Table andsalicylic 3 and their enzymatic inhibitory activities were evaluated by the previous enzyme assay method 2).Considering Further analysis Further modification of 9m was the A-ring of Schemes the xanthone core. that theA-ring substituents of activities A-ring was derived from the at starting material substituted acids, aofseries substituted compounds were synthesized according to 2relationship andsalicylic 3 and their enzymatic inhibitory activities were evaluated byconducted previous enzyme assay method (Table 2).Considering Further analysis of the inhibitory gave us athe preliminary structure–activity (SAR) the Further modification of 9m was conducted at the A-ring of the xanthone core. Considering that the substituents of A-ring was derived from the starting material substituted salicylic acids, a series of A-ring substituted compounds were synthesized according to Schemes 2 and 3 and their enzymatic inhibitory activities were evaluated by the previous enzyme assay method (Table 2). Further analysis of the inhibitory activities gave us a preliminary structure–activity relationship (SAR) of the Further modification of 9m was conducted at the A-ring of the xanthone core. Considering that the substituents of activities A-ring was derived from the starting material substituted salicylic acids, aof series of A-ring substituted compounds were synthesized according to Schemes 2relationship and 3 and their enzymatic inhibitory activities were evaluated by the previous enzyme assay method (Table 2). Further analysis of the inhibitory gave us a preliminary structure–activity (SAR) the substituents in the A-ring. All the 5or 7-substituted xanthones showed a better inhibitory activity theA-ring substituents of activities A-ring was derived from material substituted salicylic acids, athat series Further modification ofcompounds 9mevaluated was conducted atthe thestarting A-ring of the xanthone core. of substituted were synthesized according to Schemes 2relationship and 3 Considering and their enzymatic inhibitory activities were by the previous enzyme assay method (Table 2). Further analysis the inhibitory gave us a preliminary structure–activity (SAR) of the substituents in the A-ring. All the 5or 7-substituted xanthones showed a better inhibitory activity the substituents of A-ring was derived from the starting material substituted salicylic acids, a series of A-ring substituted compounds were synthesized according to Schemes 2 and 3 and their enzymatic inhibitory activities were evaluated by the previous enzyme assay method (Table 2). Further analysis the inhibitory activities gave us a preliminary structure–activity relationship (SAR) of the substituents in the A-ring. All thewere 5orasynthesized 7-substituted xanthones showed asalicylic better inhibitory activity than unsubstituted compound 15a, indicating that substitution on 2relationship A-ring might increase of A-ring substituted compounds according to Schemes and 3 and their enzymatic inhibitory activities were evaluated by previous enzyme assay method 2). Further analysis the substituents of A-ring was derived from the startingaccording material substituted acids, a series thethe gave us preliminary structure–activity (SAR) of the substituents in the activities A-ring. All thewere 5orthe 7-substituted xanthones showed a(Table better inhibitory activity than theinhibitory unsubstituted compound 15a, indicating that substitution on 2relationship A-ring might increase the of A-ring substituted compounds synthesized to Schemes and 3 and their enzymatic inhibitory activities were evaluated by the previous enzyme assay method (Table 2). Further analysis the inhibitory activities gave us a preliminary structure–activity (SAR) of the substituents in the A-ring. All the 5or 7-substituted xanthones showed a better inhibitory activity than theinhibitory unsubstituted compound 15a, indicating that substitution on A-ring might increase the interaction ofinthe inhibitors with PGAM1. Furthermore, the inhibitory effect of (7-OMe) and inhibitory activities werecompound evaluated by previous according enzyme assay method (Table 2). Further analysis of the activities gave us athe preliminary structure–activity relationship of 15e the substituents the A-ring. All the 5or 7-substituted xanthones a(Table better inhibitory activity of A-ring substituted compounds were synthesized toshowed Schemes 2 15d and 3(SAR) and their than unsubstituted 15a, indicating that substitution on A-ring might increase interaction ofinthe with PGAM1. Furthermore, the inhibitory effect of 15d (7-OMe) and inhibitory activities were evaluated by previous enzyme assay method 2). Further analysis of thethe inhibitory activities gave us a 7-substituted preliminary structure–activity relationship (SAR) of 15e the substituents theinhibitors A-ring. All the 5orthe xanthones showed a The better inhibitory activity than the unsubstituted compound 15a, indicating that substitution on A-ring might increase interaction of the inhibitors with PGAM1. Furthermore, the inhibitory effect of 15d (7-OMe) and 15e (7-Me) were slightly better than 15b (5-OMe) and 15c (5-Me), respectively. 7-chloro substituted of the inhibitory activities gave us preliminary structure–activity relationship (SAR) of2). substituents the A-ring. All the 5ora 7-substituted xanthones showed a better inhibitory activity than the unsubstituted compound 15a, indicating that substitution on A-ring might increase the interaction ofin the inhibitors with PGAM1. Furthermore, the inhibitory effect of method 15d (7-OMe) and 15e enzymatic inhibitory activities were evaluated byand the15c previous enzyme assay (Table (7-Me) were slightly better than 15b (5-OMe) (5-Me), respectively. The 7-chloro substituted of the inhibitory activities gave us preliminary structure–activity relationship (SAR) of the substituents in the A-ring. the 5ora 7-substituted xanthones showed a activity, better inhibitory activity than the unsubstituted compound 15a, indicating that substitution on A-ring might increase interaction of15f the inhibitors with PGAM1. Furthermore, the inhibitory effect of 15d (7-OMe) and 15e (7-Me) were slightly better than 15b (5-OMe) and 15c (5-Me), respectively. The 7-chloro substituted compounds with ancompound ICAll 50 of 3.5 μM could significantly enhance the while its fluorsubstituents in the A-ring. All the 5or 7-substituted xanthones showed a better inhibitory activity than the unsubstituted 15a, indicating that substitution on A-ring might increase the interaction of the inhibitors with PGAM1. Furthermore, the inhibitory effect of 15d (7-OMe) and 15e (7-Me) were slightly better than 15b (5-OMe) and (5-Me), respectively. The 7-chloro substituted Furthersubstituents analysis of the inhibitory gave ussignificantly a15c preliminary structure–activity relationship compounds 15f with ancompound IC 50 activities of 3.5 μM7-substituted could enhance theA-ring activity, while itsand fluorin the A-ring. All the 5-15a, or xanthones showed a better inhibitory activity than the unsubstituted indicating that substitution on might increase the interaction of the inhibitors with PGAM1. Furthermore, the inhibitory effect of 15d (7-OMe) 15e (7-Me) were slightly better than 15b (5-OMe) and 15c (5-Me), respectively. The 7-chloro substituted compounds 15f with ancompound IC 50 of 3.5 μM could significantly enhance theA-ring activity, while its fluorsubstituted homologs 15g (7-F) was as active as the unsubstituted compounds 15a. Comparing 15d than the unsubstituted 15a, indicating that substitution on might increase the interaction of thewith inhibitors with PGAM1. Furthermore, the inhibitory effect of 15d (7-OMe) and 15e (7-Me) were slightly better than 15b (5-OMe) and 15c (5-Me), respectively. The 7-chloro substituted compounds 15f an IC 50 of 3.5 μM could significantly enhance the activity, while its fluorsubstituted homologs 15g (7-F) was as active as the unsubstituted compounds 15a. Comparing 15d (SAR) of the substituents in the A-ring. All the 5or 7-substituted xanthones showed a better inhibitory than the unsubstituted compound 15a, indicating that substitution on A-ring might increase the interaction of the inhibitors with PGAM1. Furthermore, the inhibitory effect of 15d (7-OMe) and 15e (7-Me) were slightly better than 15b (5-OMe) and 15c (5-Me), respectively. The 7-chloro substituted compounds 15f with an IC 50 of 3.5 μM could significantly enhance the activity, while its fluorsubstituted homologs 15g (7-F) was as active as the unsubstituted compounds 15a. Comparing 15d (IC 50 = 4.6 μM) with 15e (IC 50 = 8.0 μM), we found that introducing the oxygen atom was beneficial interaction of the inhibitors with PGAM1. Furthermore, the inhibitory effect of 15d (7-OMe) and 15e were slightly better than 15b (5-OMe) and 15c (5-Me), respectively. The 15a. 7-chloro compounds 15f with an(IC IC 50 =of 3.5 μM could significantly enhance the activity, while its fluorsubstituted homologs 15g (7-F) was as active as the unsubstituted compounds Comparing 15d (IC 50 =the 4.6 μM) with 15e 50 8.0 μM), we found that introducing theeffect oxygen atom wassubstituted beneficial activity(7-Me) than unsubstituted compound 15a, indicating that on A-ring might increase interaction of the inhibitors with PGAM1. Furthermore, thesubstitution inhibitory ofwith 15d (7-OMe) and 15e (7-Me) were slightly better than 15b (5-OMe) and 15c (5-Me), respectively. The 7-chloro compounds 15f with an(IC IC 50 =of 3.5 μM could significantly enhance the activity, while its fluorsubstituted homologs 15g (7-F) was as active as the unsubstituted compounds 15a. Comparing 15d (IC 50 = 4.6 μM) with 15e 50 8.0 μM), we found that introducing thebonds oxygen atom wassubstituted beneficial for activity. We surmised that the oxygen atom might form hydrogen protein residues. (7-Me) were slightly better than 15b (5-OMe) and 15c (5-Me), respectively. The 7-chloro substituted compounds 15f with an IC 50 =of 3.5 μM could significantly enhance the activity, while its fluorsubstituted homologs 15g (7-F) was as active as the unsubstituted compounds 15a. Comparing 15d (IC 50 = 4.6 μM) with 15e (IC 50 8.0 μM), we found that introducing the oxygen atom was beneficial for activity. We surmised that the oxygen atom might form hydrogen bonds with protein residues. the interaction of the inhibitors with PGAM1. Furthermore, the inhibitory effect of 15d (7-OMe) (7-Me) were slightly better than 15b (5-OMe) and 15c (5-Me), respectively. The 7-chloro substituted compounds 15f with an IC 50 of 3.5 μM could significantly enhance the activity, while its fluorsubstituted homologs 15g (7-F) was as active as the unsubstituted compounds 15a. Comparing 15d (IC50activity. = 4.6 μM) with 15e (IC 504.6 =the 8.0 μM), we found that introducing thebonds oxygen atom was beneficial for We surmised that oxygen atom might with protein residues. Demethylation 15d15e (IC 50 =50 μM) afforded (IC 50 form =introducing 6.4 hydrogen μM), with an exposed hydroxyl compounds 15fof with an(IC IC 50 =of 3.5 could significantly enhance the activity, while itsgroup, fluorsubstituted homologs 15g (7-F) was asμM active as15i the unsubstituted compounds 15a. Comparing 15d (IC 50 = 4.6were μM) 8.0 μM), we found that the oxygen atom was7-chloro beneficial for activity. We surmised that the oxygen atom might bonds with protein residues. Demethylation of 15d15e (IC 50 =5050 4.6 μM) afforded (IC 50form =introducing 6.4 hydrogen μM), with an exposed hydroxyl group, and 15ecompounds (7-Me) slightly better than 15b (5-OMe) and 15c (5-Me), respectively. The 15fwith with an IC 3.5 μM could significantly enhance the activity, while its fluorsubstituted homologs 15g (7-F) was as active as15i the unsubstituted compounds 15a. Comparing 15d (IC 50 = 4.6 μM) with (IC =of 8.0 μM), we found that the oxygen atom was beneficial for activity. We surmised the oxygen atom might form hydrogen bonds with protein residues. Demethylation of 15d (IC 50that = 4.6 μM) afforded 15i (IC 5015d. = 6.4 μM), with an exposed hydroxyl group, slightly decreased the inhibitory activity compared to For acetylation of the exposed hydroxyl substituted homologs 15g (7-F) was as active as the unsubstituted compounds 15a. Comparing 15d (IC 50 = 4.6 μM) with 15e (IC 50 = 8.0 μM), we found that introducing the oxygen atom was beneficial for activity. We surmised that the oxygen atom might form hydrogen bonds with protein residues. Demethylation of 15d (IC 50 = 4.6 μM) afforded 15i (IC 5015d. = 6.4 μM), with an exposed hydroxyl group, slightly decreased the inhibitory activity compared to For acetylation of the exposed hydroxyl substituted homologs 15g (7-F) was as active as the unsubstituted compounds 15a. Comparing 15d substituted compounds 15f with an ofshowed 3.5we µM could significantly enhance the activity, while its (IC 50 = of 4.6 μM) 15e (IC =IC 8.0 μM), found that the oxygen atom was beneficial for activity. We surmised that the oxygen atom might form hydrogen bonds with protein residues. Demethylation of 15d (IC 50 =50 4.6 μM) afforded 15i (IC 5015d. =introducing 6.4 μM), with an exposed hydroxyl group, 50 slightly decreased the inhibitory activity compared to For acetylation of the exposed hydroxyl group 15i to with afford 15j, the latter significantly enhanced inhibitory activity. In addition, (IC 50 = 4.6 μM) with 15e (IC 50 = 8.0 μM), we found that introducing the oxygen atom was beneficial for activity. We surmised that the oxygen atom might form hydrogen bonds with protein residues. Demethylation of 15d (IC 50 = 4.6 μM) afforded 15i (IC 50 = 6.4 μM), with an exposed hydroxyl group, slightly decreased the inhibitory activity compared to 15d. For acetylation of with the exposed hydroxyl group 15i to with afford 15j, the showed significantly enhanced inhibitory activity. In beneficial addition, (IC 50 = of 4.6 μM) 15e (IC =latter 8.0 μM), we found that introducing the oxygen atom was for activity. We surmised that the oxygen atom might form hydrogen bonds protein residues. fluor-substituted homologs 15g (7-F) was as active as15i the unsubstituted compounds 15a. Comparing Demethylation of 15d (IC 50 =50 4.6 μM) afforded 50 = 6.4 μM), with an exposed hydroxyl group, slightly decreased the inhibitory activity compared to 15d. For acetylation ofpotent the exposed hydroxyl group of to of afford 15j, the latter showed significantly enhanced inhibitory activity. In hydroxyl addition, the 7-NO 215i -substituted compound 15h, with an IC 50(IC of 2.1 μM, was the most inhibitor among for activity. We surmised the oxygen atom might form hydrogen bonds with protein residues. Demethylation 15d (IC 50that = 4.6 μM) afforded 15i (IC 502.1 = 6.4 μM), with an exposed hydroxyl group, slightly decreased the inhibitory activity compared to 15d. For acetylation of the exposed group of 15i to afford 15j, the latter showed significantly enhanced inhibitory activity. In addition, the 7-NO 2 -substituted compound 15h, with an IC 50 of μM, was the most potent inhibitor among for activity. We surmised that the oxygen atom might form hydrogen bonds with protein residues. Demethylation of 15d (IC 50 = 4.6 μM) afforded 15i (IC 50 = 6.4 μM), with an exposed hydroxyl group, slightly decreased the inhibitory activity compared to 15d. For acetylation of the exposed hydroxyl 15d (IC50 = 4.6 µM) with 15e (IC = 8.0 µM), we found that introducing the oxygen atom was beneficial group of 15i to afford 15j, the latter showed significantly enhanced inhibitory activity. In addition, 7-NO 2-substituted compound 15h, with an IC50(IC of 2.1 μM, was the most potent inhibitor among 50 the A-ring substituted xanthones. Demethylation of 15d 50the = 4.6 μM) afforded 15i = 6.4 μM), with an exposed hydroxyl group, slightly the (IC inhibitory activity compared to502.1 15d. For acetylation ofpotent the exposed group ofdecreased to of afford 15j, latter showed significantly enhanced inhibitory activity. In hydroxyl addition, 7-NO 215i -substituted compound 15h, with an IC50(IC of μM, was the most inhibitor among the A-ring substituted xanthones. Demethylation 15d (IC 50 the = oxygen 4.6 μM) afforded 15i =hydrogen 6.4 μM), with an exposed hydroxyl group, slightly the inhibitory activity compared to502.1 15d. For acetylation ofpotent the exposed hydroxyl group ofdecreased 15i to afford 15j, latter showed significantly enhanced inhibitory activity. In addition, 7-NO 2surmised -substituted compound 15h, with an IC 50form of μM, was the most inhibitor among for activity. We that the atom might bonds with protein residues. the A-ring substituted xanthones. slightly the compound inhibitory activity compared to 2.1 15d. Forwas acetylation ofpotent the exposed hydroxyl group ofdecreased to afford 15j, the latter showed significantly enhanced inhibitory activity. In addition, the 7-NO 215i -substituted 15h, with an IC 50 of μM, the most inhibitor among A-ring substituted xanthones. slightly the=inhibitory activity compared to6.4 15d. Forwas acetylation ofpotent the exposed group ofdecreased to afford 15j, showed significantly enhanced inhibitory activity. In hydroxyl addition, the A-ring 7-NO 215i -substituted compound 15h, with an IC50 = of 2.1 μM, the most inhibitor among substituted xanthones. Demethylation 15d 4.6 the µM)latter afforded 15ian (IC µM), with an exposed hydroxyl group, 50 compound group ofof to (IC afford 15j, the latter showed significantly enhanced inhibitory activity. In addition, the 7-NO 215i -substituted 15h, with IC50 50 of 2.1 μM, was the most potent inhibitor among A-ring substituted xanthones. group of 15i to afford 15j, the latter showed significantly enhanced inhibitory activity. In addition, the 7-NO 2-substituted compound 15h, with an IC 50 of 2.1 μM, was the most potent inhibitor among A-ring substituted xanthones. slightlythe decreased the inhibitory activity compared to 5015d. ForμM, acetylation of the exposed hydroxyl 7-NO2-substituted compound 15h, with an IC of 2.1 was the most potent inhibitor among A-ring substituted xanthones. the 7-NO 2-substituted compound 15h, with an IC50 of 2.1 μM, was the most potent inhibitor among A-ring substituted xanthones. group of to afford 15j, the latter showed significantly enhanced inhibitory activity. In addition, the15i A-ring substituted xanthones. the A-ring substituted xanthones.
Molecules 2018, 23, 1396
5 of 15
the 7-NO2 -substituted compound 15h, with an IC50 of 2.1 µM, was the most potent inhibitor among the A-ring2018, substituted xanthones. Molecules 23, x FOR PEER REVIEW 5 of 14 Molecules 2018, 23, x FOR PEER REVIEW Molecules 2018, 23,Ox FOR PEER REVIEW O O
R1
OH
OH OH R OH 10a-10h 1 a R OH 10a-10h OH OH a 10a-10h a OH OH HO OH
O
1
HO HO
2 2
5 of 14 5 of 14
2 OH OH
R1 R1 R1
O O
O
OH b
OH OH
R1
b OH b
O R1 R1 R1
O O O O
c
R1 1
R R1
OH OH OH
O
OH OH
O OTf c c 12a-12h O OTf O OTf 12a-12h 12a-12h
R1 R1
O 11a-11h O OH 11a-11h O OH 11a-11h
O O
O
OH
O S ON O S OH N O S 15a-15h H N H O 15a-15h
e e e
R1 R1 R1
O O
OH OH OH
O N 13a-13h O N Ph Ph 13a-13h O N Ph Ph d 13a-13h Ph Ph d d O OH O O
OH OH
O NH2 14a-14h O NH2 14a-14h NH2 O 14a-14h
Scheme 2. Reagents and conditions: (a) Eaton’s reagent (7.7 wt % P2O5 solution in MeSO3H), 80 °C; ◦ C; 15a-15h Scheme 2. and conditions: (a) reagent (7.7 solution in in MeSO MeSO33H), H), 80 80 °C Scheme 2. Reagents Reagents conditions: (a) Eaton’s Eaton’s reagentdiphenylmethanimine, (7.7 wt wt % % PP22O O55 solution ; (b) Tf2O, pyridine, and DCM, 0 °C; (c) Pd(OAc) 2, BINAP, Cs2CO 3, dioxane, reflux; ◦ Scheme Reagents and conditions: (a) Eaton’s reagent (7.7 wt % P2O5 solution in 3MeSO 3H), 80 °C; (b) Tf O,2.pyridine, DCM, 0 °C C;; (c) (c) Pd(OAc) Pd(OAc) , BINAP, diphenylmethanimine, Cs CO , dioxane, reflux; (b)(d) Tf2HCl, 2O, pyridine, 22, BINAP, diphenylmethanimine, Cs22CO 3, dioxane, reflux; THF, H2DCM, O, r.t.; 0(e) [1,1′-biphenyl]-4-sulfonyl chloride, pyridine, r.t. (b) Tf2O,THF, pyridine, DCM, 0 °C;0 -biphenyl]-4-sulfonyl (c) Pd(OAc)2, BINAP, diphenylmethanimine, Cs2CO3, dioxane, reflux; (d) (d) HCl, HCl, THF, H H22O, O, r.t.; r.t.; (e) (e) [1,1 [1,1′-biphenyl]-4-sulfonyl chloride, chloride, pyridine, pyridine, r.t. r.t. (d) HCl, THF, H2O, r.t.; (e) [1,1′-biphenyl]-4-sulfonyl chloride, pyridine, r.t.
Scheme 3. Reagents and conditions: (a) BBr3, DCM, 0 °C; (b) Ac2O, NEt3, DCM. Scheme 3. Reagents and conditions: (a) BBr3, DCM, 0 °C; (b) Ac2O, NEt3, DCM. Scheme 3. 3. Reagents Reagents and and conditions: conditions: (a) (a) BBr BBr3, DCM, 0 ◦°C Scheme C;; (b) Ac22O, NEt33,, DCM. DCM. Table 2. Inhibitory activities of3 15a–15k against PGAM1. Table 2. Inhibitory activities of 15a–15k against PGAM1. Table 2. Inhibitory activities of 15a–15k against PGAM1.
Entry Compounds R1 IC50 (μM) * 1 Entry Compounds R IC 505.5 (μM) * 9m 1 8-OH ± 1.1 Entry Compounds R1R1 IC 50 (μM) ** Compounds IC (µM) 50 9m 1 2 Entry 8-OH 5.5 ± 1.1 15a H 14.3 ± 0.8 9m 8-OH 5.5 ±± 1 8-OH 5.56.5 1.1 15a 21 3 H 14.3 ±1.1 15b9m 5-OMe ±0.8 1.3 15a 2 H 14.3 ± 0.8 2 15a H 14.3 ± 15b 3 4 5-OMe 6.58.6 ± 1.3 15c 5-Me ±0.8 3.1 3 5-OMe 6.54.6 15b 5-OMe 6.5 1.3 15c 43 5 5-Me 8.6 ±±±3.1 15d15b 7-OMe ±1.3 0.8 4 15c 5-Me 8.6 ±±3.1 3.1 15c 4 5-Me 8.6 15d 5 6 7-OMe 4.68.0 ± 0.8 15e 7-Me ± 2.1 5 15d 7-OMe 4.6 ±±0.8 0.8 15d 7-OMe 4.6 15e 65 7 7-Me 8.0 ±±2.1 15f15e 7-Cl ±2.1 1.0 6 7-Me 8.03.5 15e 7-Me 8.0 2.1 15f 76 8 7-Cl 3.5 ±±±1.0 15g15f 7-F 13.7 ± 5.5 7 7-Cl 3.5 1.0 15f 7-Cl 3.52.1 ±±±1.0 15g 87 9 7-F 13.7 8 7-F 2 13.7 5.5 15h15g ±5.5 0.2 7-NO 15g 8 7-F 13.7 ±0.2 9 15h 7-NO 2.1 0.2 15h 2 2 2.16.4 ±± 9 10 7-NO 15i 7-OH ±5.5 1.0 15h 2 2.1 0.2 9 11 10 7-NO 7-OH 6.42.7 15i15j 15i 10 7-OH 6.4 ±±±1.0 7-OAc ±1.0 0.5 7-OAc 2.7 ±±1.0 0.5 15i 15j 1012 11 7-OH 6.4 15j 11 2.7 ± 0.5 PGMI-004A 7-OAc 13 ± 0.1 12 PGMI-004A 13 ± 0.1 15j 11 7-OAc 2.7 ± 0.5 12 * All IC50 values are PGMI-004A 13 ± 0.1 reported as means ± SD from three determinations. 12 PGMI-004A 13 ± 0.1 All IC values are as as means ± SD fromfrom threethree determinations. ** All IC50 50 values arereported reported means ± SD determinations. * All IC50 values are reported as means ± SD from three determinations.
In order to evaluate the specificity of these N-xanthone benzenesulfonamides against PGAM1, In order to evaluate of these benzenesulfonamides against PGAM1, 13 compounds with IC50the lessspecificity than 10 μM were N-xanthone selected to measure the inhibitory activities against 3 In order towith evaluate the than specificity ofwere theseselected N-xanthone benzenesulfonamides against against PGAM1, 13downstream compounds IC50 less 10 μMof towas measure the inhibitory activities 3 In order toenzymes. evaluate the specificity these N-xanthone benzenesulfonamides against The counter-screen assay [14,22] then performed to exclude thePGAM1, off-target 13 compounds with IC50 less than 10 μM were selected to measure the inhibitory activities against 3 downstream enzymes. The counter-screen assay [14,22] was then performed to exclude the off-target 13inhibition compounds with ICother 10 µMinwere to measure the inhibitory against against enzymes theselected in vitro assay (Table 3). Allactivities 13 N-xanthone 50 less than downstreamagainst enzymes. The counter-screen assay [14,22] wasassay then performed the off-target inhibition other enzymes the inhibitory in vitro 3).to exclude All 13 N-xanthone 3 benzenesulfonamides downstream enzymes. The counter-screen assay [14,22] then performed to exclude the showed a muchinlower effectwas on (Table the counter-screen assay compared inhibition against other enzymes in theinhibitory in vitro assay (Table 3). All assay 13 N-xanthone benzenesulfonamides showed a much lower effect on the counter-screen compared off-target inhibition against other enzymes in the in vitro assay (Table 3). All 13 N-xanthone to PGMI-004A, which suggests that the N-xanthone benzenesulfonamides selectively inhibit PGAM1 benzenesulfonamides showed athat much lower inhibitory effect on the counter-screen assay compared to PGMI-004A, which showed suggests thelower N-xanthone benzenesulfonamides selectively inhibit PGAM1 benzenesulfonamides a much inhibitory effect on the counter-screen assay compared to in the multiple-enzymes coupled assay system. to PGMI-004A, which suggests that the N-xanthone benzenesulfonamides selectively inhibit PGAM1 in the multiple-enzymes coupled assay system. in the multiple-enzymes coupled assay system.
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PGMI-004A, which suggests that the N-xanthone benzenesulfonamides selectively inhibit PGAM1 in the multiple-enzymes coupled system. assay of the representative compounds. Table 3. Theassay counter-screen Counter-Screen Assay (5 μM) * EntryTable Compounds PGAM1 assay IC50 (μM) 3. The counter-screen of the representative compounds. 9h 1 6.4 ± 0.4 12% Entry Compounds PGAM1 IC (µM) Counter-Screen Assay (5 µM) * 9k 2 8.4 ± 2.7 50 12% 1 3 6.4 ± 0.4 9l 9h 5.9 ± 1.2 15% 12% 2 9k 8.4 ± 2.7 12% 9m 4 5.5 ± 1.1 9% 15% 3 9l 5.9 ± 1.2 9n 9m 6.0 ± 0.6 13% 9% 4 5 5.5 ± 1.1 5 6 6.0 ± 0.6 15b 9n 6.5 ± 1.3 13% 13% 6 7 15b 6.5 ± 1.3 15c 8.6 ± 3.1 7% 13% 7 15c 8.6 ± 3.1 7% 15d 8 4.6 ± 0.8 6% 8 15d 4.6 ± 0.8 6% 15e15e 8.0 ± 2.1 7% 7% 9 9 8.0 ± 2.1 15f15f 3.5 ± 1.0 8% 8% 1010 3.5 ± 1.0 1111 2.1 ± 0.2 15h15h 2.1 ± 0.2 5% 5% 1212 15i 6.4 ± 1.0 15i 6.4 ± 1.0 10% 10% 13 15j 2.7 ± 0.5 7% 15j 13 2.7 ± 0.5 7% 38% 14 PGMI-004A 13 ± 0.1 PGMI-004A 14 13 ± 0.1 38%
* The inhibition ratios against the other three enzymes were measured at a concentration of 5 µM.
* The inhibition ratios against the other three enzymes were measured at a concentration of 5 μM.
To better understand the structure–activity relationship of the A-ring of the xanthone core, To better understand the structure–activity relationship of the A-ring of the xanthone core, 15h 15h was docked into the crystal structure of PGAM1. The interactions between 15h and PGAM1 are was docked into the crystal structure of PGAM1. The interactions between 15h and PGAM1 are shown in Figure 3. The biphenyl group occupied the hydrophobic pocket formed by P123, F22, L95, shown in Figure 3. The biphenyl group occupied the hydrophobic pocket formed by P123, F22, L95, and W115. The carbonyl and the sulfone amide group in the 15h contacted with E89 and R116 through and W115. The carbonyl and the sulfone amide group in the 15h contacted with E89 and R116 through hydrogen bonds, respectively. In addition, the 7-nitro group formed two hydrogen bonds with S23, hydrogen bonds, respectively. In addition, the 7-nitro group formed two hydrogen bonds with S23, which enhanced the intramolecular interaction between 15h and the enzyme. which enhanced the intramolecular interaction between 15h and the enzyme.
Figure 3. The docking model of 15h with PGAM1. PGAM1.
After two tworounds roundsofof optimization, enzymatic inhibitory activity 50) against PGAM1 of After optimization, thethe enzymatic inhibitory activity (IC50(IC ) against PGAM1 of these these N-xanthone benzenesulfonamides was increased from 20 µM. μM Then, to 2.1 μM.representative Then, some N-xanthone benzenesulfonamides was increased from 20 µM to 2.1 some representative compounds were picked out to evaluate their antiproliferative activity (H1299 cell compounds were picked out to evaluate their antiproliferative activity (H1299 cell line) by the MTT line) by the MTT method (Table 4). Among them, 9h showed a relatively strong inhibitory activity method (Table 4). Among them, 9h showed a relatively strong inhibitory activity against H1299 against H1299 cell proliferation. cell proliferation.
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Molecules 2018, 23, x FOR PEER REVIEW 7 of 14 Molecules 2018, 23, x FOR PEER REVIEW of 14 Table 4. Antiproliferation activities of representative compounds. 77 of Molecules 2018, 23, x FOR PEER REVIEW 14 Molecules 2018, 23, x FOR PEER REVIEW 7 of 14 TableREVIEW 4. Antiproliferation activities of representative compounds. Molecules 2018, 23, x FOR PEER 7 of 14 TableREVIEW 4. Antiproliferation activities of representative compounds. Molecules 2018, 23, x FOR PEER 7 of 14 Table 4. Antiproliferation activities of representative compounds. Molecules 23, x FOR PEER REVIEW 7 of *14 Entry 2018, Compounds Structures IC50 (µM) Entry2018, Compounds Structures IC50 (μM) 4. Antiproliferation activities of representative compounds. Molecules 23, x FOR Table PEER REVIEW 7*of 14 Entry CompoundsTable 4. Antiproliferation activities Structures IC50 (μM) * of representative compounds. Molecules 23, x FOR Table PEER REVIEW Entry2018, Compounds Structures IC50 (μM)7*of 14 4. Antiproliferation activities of representative compounds. Entry Compounds Table 4. Antiproliferation activities Structures IC50 (μM) * of representative compounds. Entry Compounds Structures IC 50 (μM) * Table 4. Antiproliferation activities of representative compounds. 9h 1 14.1 ±14.1 1.9 ± 1.9 1 9h Entry Structures IC 50 (μM) * 9h 1 EntryCompounds 14.1 1.9 Table 4. Antiproliferation activities of representative compounds. Compounds Structures IC±±501.9 (μM) * 9h 1 14.1 Entry Compounds Structures IC 50 (μM) * 9h 1 14.1 ± 1.9 9h 1 14.1 ± 1.9 * Entry Compounds Structures IC 50 (μM) 9h 1 14.1 ± 1.9 9h 1 14.1 ± 1.9 9h 12 14.1 9k 21.1 ± 1.9 1.7 9k 9k 22 21.1 ±21.1 1.7 9h 14.1 ±± 1.7 1.9 ± 1.7 9k 21 21.1 9k 2 21.1 ± 1.7 9k 2 21.1 ± 1.7 9k 2 21.1 ± 1.7 9k 2 21.1 ± 1.7 9k 23 21.1 9l 18.5 ±± 1.7 2.3 9l 18.5 ± 1.7 2.3 9k 23 21.1 9l 9l 33 18.5 ±18.5 2.3 ± 2.3 9l 3 18.5 ± 2.3 9l 3 18.5 ± 2.3 9l 3 18.5 ± 2.3 9l 3 18.5 ± 2.3 9l 3 18.5 ± 2.3 9m 4 20.1 ± 1.4 9l 34 18.5 9m 20.1 ±± 2.3 1.4 9m 4 20.1 ± 1.4 9m9m 44 20.1 ±20.1 1.4 ± 1.4 9m 4 20.1 ± 1.4 9m 4 20.1 ± 1.4 9m 4 20.1 ± 1.4 9m 4 20.1 ± 1.4 15b 42.7 ±± 1.4 3.9 9m 45 20.1 15b 5 42.7 ± 3.9 15b 5 42.7 ± 3.9 15b 5 42.7 ± 3.9 15b15b 55 42.7 ±42.7 3.9 ± 3.9 15b 5 42.7 ± 3.9 15b 5 42.7 ± 3.9 15b 5 42.7 ± 3.9 15d 19.6 ± 2.2 15b 566 42.7 15d 19.6 ±± 3.9 2.2 15d 6 19.6 ± 2.2 15d 6 19.6 ± 2.2 15d 6 19.6 ± 2.2 15d15d 66 19.6 ±19.6 2.2 ± 2.2 15d 6 19.6 ± 2.2 15d 6 19.6 ± 2.2 15f 7 32.7 ± 0.9 15d 67 19.6 15f 32.7 ± 2.2 0.9 15f 7 32.7 ± 0.9 15f 7 32.7 ± 0.9 15f 7 32.7 ± 0.9 15f 7 32.7 ± 0.9 15f 32.7 ± 0.9 7 7 15f 32.7 ± 0.9 15f 7 32.7 ± 0.9 15h 8 33.8 ± 6.1 15f 78 32.7 15h 33.8 ± 0.9 6.1 15h 8 33.8 ± 6.1 15h 8 33.8 ± 6.1 15h 8 33.8 ± 6.1 15i 30.6 ±± 6.1 2.4 15h 89 33.8 15i 15h 9 8 30.633.8 ± 2.4± 6.1 15i 9 30.6 ±33.8 2.4 ± 6.1 15h15h 88 33.8 6.1 15i 9 30.6 ± 2.4 15i 98 30.6 15h 33.8 ± 2.4 6.1 15j 10 31.1 ±± 2.4 4.5 15i 9 30.6 15j 15i 10 9 31.130.6 ± 4.5± 2.4 15j 10 31.1 ± 2.4 4.5 15i 9 30.6 15j15i 109 31.1 ±30.6 4.5 ± 2.4 15j 15i 10 31.1 9 30.6 ± 4.5 2.4 Gemcitabine 0.193 15j 10 31.1 ± 4.5 Gemcitabine 0.193 15j 10 31.1 PGMI-004A 26.0 2.1± 4.5 Gemcitabine 0.193 15j 10 31.1 ±±± 4.5 PGMI-004A 26.0 2.1 Gemcitabine 0.193 PGMI-004A 26.0 ±31.1 2.1 Gemcitabine 0.193 15j15j 1010 31.1 4.5 ± 4.5 * All IC50 values are reported as means ± SD from three determinations. PGMI-004A 26.0 ± 2.1 Gemcitabine 0.193 * All IC50 values are reported as means ± SD from three determinations. PGMI-004A 26.0 ± 0.193 2.1 Gemcitabine * All IC50 values are reported as means ± SD from three determinations. PGMI-004A 26.0 ± 2.1 Gemcitabine 0.193 * All IC50 values are reported as means ± SD from three determinations. PGMI-004A 26.0 ± 2.1 * All IC50 values are reported as means ± SD from three determinations. 3. Experimental Section Gemcitabine 0.193 PGMI-004A 26.0 ± 2.1 * All IC50 values are reported as means ± SD from three determinations. 3. Experimental Section Gemcitabine * All IC50 values are reported as means ± SD from three determinations. 26.0 ± 2.10.193 3. Experimental Section PGMI-004A * All IC50 values are reported as means ± SD from three determinations. 3. Experimental Section
3. Experimental Section 3.1. Chemistry Experimental Procedures and Compound Characterization PGMI-004A 26.0 ± 2.1 * All IC50 values are reported as meansCharacterization ± SD from three determinations. 3. Experimental Section 3.1. Chemistry Experimental Procedures and Compound Experimental SectionProcedures and Compound Characterization 3.1.3.Chemistry Experimental * All IC values are reported as means ± SD from three determinations. 3. Experimental Section 50 3.1. Chemistry Experimental Procedures and Characterizationacid (5.0 g, 32.4 mmol) and 1,3,8-Trihydroxy-9H-xanthen-9-one (3). Compound 2,6-dihydroxybenzoic 3.1. Chemistry Experimental Procedures and Characterizationacid (5.0 g, 32.4 mmol) and 1,3,8-Trihydroxy-9H-xanthen-9-one (3). Compound 2,6-dihydroxybenzoic 3. Experimental Section 3.1. Chemistry Experimental and Compound Characterization 1,3,8-Trihydroxy-9H-xanthen-9-one (3).added 2,6-dihydroxybenzoic acid 32.4onmmol) phloroglucinol (4.09 g, 32.4Procedures mmol) were to Eaton’s reagent (20 mL)(5.0 and g, heated an 80 °Cand oil 3.1. Chemistry Experimental Procedures and Compound Characterization 1,3,8-Trihydroxy-9H-xanthen-9-one (3).added 2,6-dihydroxybenzoic acid 32.4onmmol) phloroglucinol (4.09 g, 32.4 mmol) were to Eaton’s reagent (20 mL)(5.0 and g, heated an 80 °Cand oil 3.1. Chemistry Experimental and Compound Characterization 1,3,8-Trihydroxy-9H-xanthen-9-one (3).added 2,6-dihydroxybenzoic acid 32.4 and phloroglucinol (4.09 g, 32.4toProcedures mmol) were tosolution Eaton’s reagent (20 mL)(5.0 and g, heated onmmol) an ice 80 °C oil bath for 2 h. After cooling r.t., the dark-brown was transferred slowly to crushed while 1,3,8-Trihydroxy-9H-xanthen-9-one (3). 2,6-dihydroxybenzoic acid (5.0 g, 32.4 mmol) and phloroglucinol (4.09 g, 32.4toProcedures mmol) tosolution Eaton’s reagent (20 mL) and heated on anmmol) 80 °C oiland bath for 2 h. After cooling r.t., thewere dark-brown was transferred slowly to crushed ice while 3.1. Chemistry Experimental andadded Compound Characterization 1,3,8-Trihydroxy-9H-xanthen-9-one (3). 2,6-dihydroxybenzoic acidslowly (5.0 32.4 phloroglucinol (4.09 g, 32.4 added tosolution Eaton’s reagent (20 mL) and heated onwater, an ice 80 dried, °C oil bath for 2 h. After cooling tommol) r.t., thewere dark-brown was transferred tog, crushed while being vigorously stirred. The mixture was filtered and the filter cake was washed with 1,3,8-Trihydroxy-9H-xanthen-9-one (3). 2,6-dihydroxybenzoic acid (5.0 32.4 mmol) and (4.09 g, 32.4 mmol) were added tosolution Eaton’s reagent (20 mL) and g, heated onwater, an ice 80 dried, °C oil bath for 2 h. After cooling to r.t., the dark-brown was transferred slowly to crushed while being vigorously stirred. The mixture was filtered and the filter cake was washed with 3.phloroglucinol Experimental Section phloroglucinol (4.09column g,The 32.4 mmol) were added to Eaton’s reagent (20 mL) and onice an 80and °C oil bath for 2 h. After cooling to r.t., the dark-brown solution was transferred slowly toheated crushed while being vigorously stirred. mixture was filtered and the filter cake was washed with water, dried, and purified by flash chromatography to afford xanthone 3 as a yellow solid (2.3 g, yield: 1,3,8-Trihydroxy-9H-xanthen-9-one (3). 2,6-dihydroxybenzoic acid (5.0 g, 32.4 mmol) phloroglucinol g,column 32.4 added toto Eaton’s reagent (203 was mL) and heated onwater, an ice 80 °C oil bath for 2 h. After cooling tommol) r.t., thewere dark-brown solution was transferred tosolid crushed while being vigorously stirred. The mixture was filtered and the filter cake washed with andbath purified by(4.09 flash chromatography afford xanthone as aslowly yellow (2.3 g, dried, yield: for 2 h. After cooling to r.t.,were dark-brown solution was transferred slowly to on crushed ice while 1vigorously being stirred. The mixture was filtered and the(s, filter cake was washed with water, dried, and purified by flash column chromatography to afford xanthone 3 as a yellow solid (2.3 g, yield: 29%). H NMR (400 MHz, DMSO-d 6the ) δ 11.85 (s, 1H), 11.80 1H), 11.28 (s, 1H), 7.68 (tt, J = 8.3, 2.4 Hz, phloroglucinol (4.09 g, 32.4 mmol) added to Eaton’s reagent (20 mL) and heated an 80 °C oil bath for h. After cooling to r.t., the6)dark-brown solution transferred tosolid crushed ice while 1vigorously being stirred. The mixture filtered and thewas filter cake washed with dried, and purified by flash column chromatography to afford xanthone 3 was as(s,was aslowly yellow (2.3 g,2.4 yield: 29%). H2NMR (400 MHz, DMSO-d δwas 11.85 (s,filtered 1H), 11.80 (s, 1H), 11.28 1H), 7.68 (tt,with J water, =(2.3 8.3, Hz, 3.1. Chemistry Experimental Procedures and Compound Characterization 1 being vigorously stirred. The mixture was and the filter cake washed water, dried, and purified by flash column chromatography to afford xanthone 3 as a yellow solid g, yield: 29%). H2NMR MHz, DMSO-d 6)dark-brown δ(ddd, 11.85J(s, 1H), 11.80 (s, 1H), 11.28 (s, 1H), 7.68 (tt,1H), J = 8.3, 2.4 Hz, 1H),purified 7.00 (dt, J =(400 8.3, 3.3 Hz, 1H), 6.79 = 8.4, 3.0, 1.8 Hz, 1H), 6.39 (d, J = 1.9 Hz, 6.23 (d, J= bath for h. After cooling to r.t., the solution transferred to crushed ice while 1vigorously being stirred. The mixture was and thewas filter cake washed with dried, and flash chromatography to afford xanthone 3 was as(s, aslowly yellow solid (2.3 g,2.4 yield: 29%). Hpurified NMR MHz, DMSO-d 6) δ(ddd, 11.85filtered 1H), 11.80 (s, 1H), 11.28 1H), 7.68 (tt,1H), J water, = 8.3, 1H), 7.00 (dt, by J =(400 8.3, 3.3column Hz, 1H), 6.79 J(s, = 8.4, 3.0, 1.8 Hz, 1H), 6.39 (d, J = 1.9 Hz, 6.23 (d, J= 1H and by flash column chromatography to afford xanthone 3(d, as yellow solid (2.3 g,Hz, yield: 29%). NMR MHz, DMSO-d 6) δ 11.85filtered 1H),3.0, 11.80 (s, 1H), 11.28 (s, 1H), 7.68 (tt,1H), J water, = 8.3, 2.4 Hz, 1H), 7.00 (dt, Jby =(400 8.3, 3.3column Hz, 1H), 6.79 (ddd, J(s, = 8.4, 1.8 Hz, 1H), 6.39 J a= 1.9 Hz, 6.23 (d, J= 1.5 Hz, 1H). being vigorously stirred. The mixture was and the filter cake was washed with dried, 1 and purified flash chromatography to afford xanthone 3 as a yellow solid (2.3 g, yield: 1,3,8-Trihydroxy-9H-xanthen-9-one (3). 2,6-dihydroxybenzoic acid g, 2.4 32.4 mmol) and 29%). H1H). NMR MHz, DMSO-d 6) δ(ddd, 11.85 1H), 11.80 (s, 11.28 (s, 7.68 (tt,(5.0 J = 8.3, Hz, 1H), 7.00 J =(400 8.3, 3.3MHz, Hz, 1H), 6.79 J(s, = 8.4, 3.0, 1.8 Hz,1H), (d,1H), J =1H), 1.9 Hz, 1H), (d, J =Hz, 1.5 Hz, 1(dt, H NMR (400 DMSO-d 6) δ 11.85 (s, 11.80 (s,1H), 1H),6.39 (s, 7.68 (tt, J6.23 = g, 8.3, 2.4 1H), 7.00 (dt, J = 8.3, 3.3 Hz, 1H), 6.79 (ddd, J = 8.4, 3.0, 1.8 Hz, 1H), 6.39 (d, J = 1.9 Hz, 1H), 6.23 (d, J= 1.5 Hz, 1H). and29%). purified flash chromatography to1H), afford xanthone 311.28 as(s, a1H), solid (2.3 yield: 1H 29%). NMR MHz, DMSO-d 6) δ 11.85 1H), 11.80 (s, 1H), 7.68 (tt,1H), J =and 8.3, 2.4 Hz, 1H), 7.00 (dt, Jby =(400 8.3, 3.3column Hz, 1H), 6.79 (ddd, J (s, = 8.4, 3.0, 1.8 Hz, 1H),11.28 6.39 (d, Jyellow = 1.9 Hz, 6.23 (d, J = on an 80 ◦ C 1.5 Hz, 1H). phloroglucinol (4.09 g, 32.4 mmol) were added to Eaton’s reagent (20 mL) heated 1H), 7.00 (dt, J = 8.3, 3.3DMSO-d Hz, 1H),6)6.79 (ddd, J1H), = 8.4,11.80 3.0, 1.8 1H), Hz, 1H), 6.39 (d, J 7.68 = 1.9(tt, Hz, 6.23Hz, (d, J = 1.5 Hz,1H1H). 29%). NMR 11.85 J =1H), 8.3, 1H), 7.00 (dt, J =(400 8.3, MHz, 3.3 Hz, 1H), 6.79 δ(ddd, J (s, = 8.4, 3.0, 1.8 (s, Hz, 1H),11.28 6.39 (s, (d,1H), J = 1.9 Hz, 1H), 6.232.4 (d, J = Hz, 1H). oil1.5 bath for 2 h. After cooling to r.t., the dark-brown solution was transferred slowly to crushed 1.5 Hz, 1H). 1H),Hz, 7.00 (dt, J = 8.3, 3.3 Hz, 1H), 6.79 (ddd, J = 8.4, 3.0, 1.8 Hz, 1H), 6.39 (d, J = 1.9 Hz, 1H), 6.23 (d, J = 1.5 1H). ice being vigorously stirred. The mixture was filtered and the filter cake was washed with 1.5while Hz, 1H).
water, dried, and purified by flash column chromatography to afford xanthone 3 as a yellow solid (2.3 g, yield: 29%). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.85 (s, 1H), 11.80 (s, 1H), 11.28 (s, 1H),
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7.68 (tt, J = 8.3, 2.4 Hz, 1H), 7.00 (dt, J = 8.3, 3.3 Hz, 1H), 6.79 (ddd, J = 8.4, 3.0, 1.8 Hz, 1H), 6.39 (d, J = 1.9 Hz, 1H), 6.23 (d, J = 1.5 Hz, 1H). 1,8-Dihydroxy-9-oxo-9H-xanthen-3-yl-trifluoromethanesulfonate (4). To a solution of compound 3 (2.0 g, 8.2 mmol) and NEt3 (1.4 mL, 9.8 mmol) in dry DCM (20 mL), Tf2 O (1.5 mL, 9.0 mmol) was added slowly at 0 ◦ C. The reaction was completed in 2 h, shown by TLC analysis, and was then diluted with DCM (20 mL) and hydrochloric acid (1 mol/L, 40 mL). The organic phase was separated, washed with brine, dried over Na2 SO4 , filtered, and concentrated in vacuo successively. The residue was purified by flash column chromatography to afford triflate 4 as a yellow solid (1.5 g, yield: 48%). 1 H NMR (400 MHz, DMSO) δ 12.15 (s, 1H), 11.58 (s, 1H), 7.78 (t, J = 8.4 Hz, 1H), 7.36 (d, J = 2.2 Hz, 1H), 7.08 (d, J = 8.4 Hz, 1H), 7.04 (d, J = 2.2 Hz, 1H), 6.88 (d, J = 8.3 Hz, 1H). 9-Oxo-3-(((trifluoromethyl)sulfonyl)oxy)-9H-xanthene-1,8-diyl bis(2,2-dimethylpropanoate) (5). To a solution of triflate 4 (1.0 g, 2.66 mmol) in dry THF (20 mL), 60% NaH (532 mg, 13.3 mmol) was added portionwise at 0 ◦ C. After 20 min, pivaloyl chloride (0.98 mL, 8.0 mmol) was added slowly. The reaction was kept at 0 ◦ C for another 1 h and then diluted with ethyl acetate (40 mL) and quenched with sat. NH4 Cl solution (60 mL). The organic phase was separated, washed with brine, dried over Na2 SO4 , filtered, and concentrated in vacuo successively. The residue was recrystallized from methanol/ethyl acetate to afford the dipivalate 5 as a white solid (1.3 g, yield: 90%). 3-((Diphenylmethylene)amino)-9-oxo-9H-xanthene-1,8-diyl bis(2,2-dimethylpropanoate) (6). The dipivalate 5 (1.0 g, 1.84 mmol), benzophenone imine (666 mg, 3.67 mmol), BINAP (172 mg, 0.275 mmol), Pd(OAc)2 (41 mg, 0.184 mmol), and Cs2 CO3 (898 mg, 2.75 mmol) were suspended in dioxane (30 mL) under argon and the resulting mixture was heated to reflux for 12 h. After cooling to r.t., the reaction mixture was diluted with ethyl acetate (50 mL), filtered through a celite bed and concentrated in vacuo successively. The residue was purified by flash column chromatography to afford the desired product 6 as a yellow solid (821 mg, yield: 77%). 3-Amino-9-oxo-9H-xanthene-1,8-diyl bis(2,2-dimethylpropanoate) (7). To a solution of compound 6 ( mg, 0.372 mmol) in THF (10 mL), 4 M hydrochloric acid (4 mL) was added. After 30 min, the reaction mixture was diluted with ethyl acetate (10 mL) and quenched with saturated aqueous sodium bicarbonate solution (15 mL). The organic phase was separated, washed with brine, dried over sodium sulfate, and concentrated in vacuo successively. The residue was crystallized in hexane/ethyl acetate to afford the desired amine 7 as a yellow solid (85 mg, yield: 80%). 1 H NMR (400 MHz, DMSO) δ 7.73–7.69 (m, 1H), 7.42 (dd, J = 8.5, 1.0 Hz, 1H), 6.95 (dd, J = 7.9, 1.0 Hz, 1H), 6.65 (brs, 2H), 6.41 (d, J = 2.1 Hz, 1H), 6.21 (d, J = 2.1 Hz, 1H), 1.35 (s, 9H), 1.33 (s, 9H). The general synthetic procedure of compounds 9a–9n. To a solution of amine 7 (41 mg, 0.1 mmol) in dry pyridine (2 mL), substituted benzenesulfonyl chloride (1.5–2 eq.) was added. The reaction mixture was kept at r.t. overnight and poured into a mixture of 1 M hydrochloric acid (10 mL) and ethyl acetate (10 mL) while being vigorously stirred. The organic phase was separated and concentrated in vacuo. The residue was dissolved in a mixture of methanol (10 mL) and 5 M sodium hydroxide solution (5 mL) and kept at r.t. for 1 h. The mixture was concentrated in vacuo to remove the methanol and was diluted with water (3 mL) and filtered. The clear water phase was washed with ethyl acetate (3 mL × 2) and then concentrated hydrochloric acid was added dropwise until pH = 4. The mixture was filtered to afford the desired sulfonamide 9a–9n as yellow solids (yield: 30–80%). N-(1,8-Dihydroxy-9-oxo-9H-xanthen-3-yl)-1-phenylmethanesulfonamide (9a). Yield: 53%. ◦ 1 Yellow solid. M.p = 210–212 C. Rf = 0.35 (Petroleum ether: Acetone = 4:1). H NMR (400 MHz, DMSO-d6 ) δ 11.79 (s, 1H), 11.77 (s, 1H), 10.75 (s, 1H), 7.73 (t, J = 8.4 Hz, 1H), 7.39–7.33 (m, 3H), 7.32–7.26 (m, 2H), 7.07 (d, J = 8.4 Hz, 1H), 6.83 (d, J = 8.3 Hz, 1H), 6.78 (s, 1H), 6.56 (s, 1H), 4.71 (s, 2H). MS (ESI− ) m/z 396.0 (M − H)− . HRMS (ESI− ): Calcd. for C20 H14 NO6 S− [M − H]− m/z: 396.0547, found: 396.0559.
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N-(1,8-Dihydroxy-9-oxo-9H-xanthen-3-yl)benzenesulfonamide (9b). Yield: 68%. Yellow solid. M.p = 260–263 ◦ C. Rf = 0.33 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.71 (s, 1H), 11.69 (s, 1H), 11.40 (s, 1H), 7.92 (d, J = 7.9 Hz, 2H), 7.73–7.56 (m, 4H), 7.02 (d, J = 8.6 Hz, 1H), 6.79 (d, J = 8.3 Hz, 1H), 6.72 (s, 1H), 6.53 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 183.76, 161.25, 160.24, 156.69, 155.58, 146.64, 139.10, 137.64, 133.67, 129.74, 126.69, 110.81, 107.30, 107.26, 103.46, 99.41, 95.48. MS (ESI− ) m/z 382.0 (M − H)− . HRMS (ESI− ): Calcd. for C19 H12 NO6 S− [M − H]− m/z: 382.0391, found: 382.0400. N-(1,8-Dihydroxy-9-oxo-9H-xanthen-3-yl)-4-fluorobenzenesulfonamide (9c). Yield: 64%. Yellow solid. M.p = 241–242 ◦ C. Rf = 0.30 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.72 (s, 1H), 11.69 (s, 1H), 11.47 (s, 1H), 8.00 (dd, J = 8.7, 5.0 Hz, 2H), 7.69 (t, J = 8.3 Hz, 1H), 7.49 (t, J = 8.6 Hz, 2H), 7.02 (d, J = 8.4 Hz, 1H), 6.80 (d, J = 8.2 Hz, 1H), 6.73 (s, 1H), 6.54 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 183.80, 164.71 (d, J = 252.8 Hz), 161.28, 160.25, 156.71, 155.58, 146.35, 137.68, 135.40, 129.91 (d, J = 9.6 Hz), 117.02 (d, J = 22.9 Hz), 110.84, 107.32, 107.26, 103.61, 99.50, 95.59. MS (ESI− ) m/z 399.9 (M − H)− . HRMS (ESI− ): Calcd. for C19 H11 FNO6 S− [M − H]− m/z: 400.0297, found: 400.0302. Yield: 75%. N-(1,8-Dihydroxy-9-oxo-9H-xanthen-3-yl)-2-fluorobenzenesulfonamide (9d). Yellow solid. M.p = 244–246 ◦ C. Rf = 0.30 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.73 (s, 1H), 11.69 (s, 1H), 11.55 (s, 1H), 7.76–7.58 (m, 4H), 7.02 (d, J = 8.4 Hz, 1H), 6.80 (d, J = 8.3 Hz, 1H), 6.75 (s, 1H), 6.55 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 183.80, 161.27, 160.23, 158.20 (d, J = 255.2 Hz), 156.68, 155.59, 137.66, 136.79 (d, J = 7.8 Hz), 130.51, 126.51 (d, J = 13.2 Hz), 125.40, 117.68 (d, J = 20.1 Hz), 110.83, 107.33, 107.27, 103.54, 99.15, 95.27, 90.84. MS (ESI− ) m/z 399.9 (M − H)− . HRMS (ESI− ): Calcd. for C19 H11 FNO6 S− [M − H]− m/z: 400.0297, found: 400.0307. 4-Chloro-N-(1,8-dihydroxy-9-oxo-9H-xanthen-3-yl)benzenesulfonamide (9e). Yield: 56%. Yellow solid. M.p = 257–258 ◦ C. Rf = 0.37 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.72 (s, 1H), 11.68 (s, 1H), 11.47 (s, 1H), 7.92 (d, J = 8.2 Hz, 2H), 7.76–7.60 (m, 3H), 7.01 (d, J = 8.4 Hz, 1H), 6.79 (d, J = 8.4 Hz, 1H), 6.72 (s, 1H), 6.53 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 183.81, 161.30, 160.25, 156.72, 155.58, 146.23, 138.61, 137.87, 137.69, 129.93, 128.68, 110.85, 107.33, 107.26, 103.68, 99.56, 95.65. MS (ESI− ) m/z 416.0 (M − H)− . HRMS (ESI− ): Calcd. for C19 H11 ClNO6 S− [M − H]− m/z: 416.0001, found: 416.0011. N-(1,8-Dihydroxy-9-oxo-9H-xanthen-3-yl)-4-iodobenzenesulfonamide (9f). Yield: 69%. Yellow solid. M.p = 269–272 ◦ C. Rf = 0.32 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.79 (s, 1H), 11.69 (s, 1H), 7.99 (d, J = 8.1 Hz, 2H), 7.76–7.56 (m, 3H), 7.00 (d, J = 8.1 Hz, 1H), 6.78 (d, J = 8.1 Hz, 1H), 6.64 (s, 1H), 6.44 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 183.34, 161.13, 160.23, 156.71, 155.55, 138.78, 138.37, 137.39, 128.97, 128.25, 110.68, 107.20, 102.77, 101.28, 100.05, 95.92. MS (ESI− ) m/z 508.0 (M − H)− . HRMS (ESI− ): Calcd. for C19 H11 INO6 S− [M − H]− m/z: 507.9357, found: 507.9376. N-(1,8-Dihydroxy-9-oxo-9H-xanthen-3-yl)-3,5-difluorobenzenesulfonamide (9g). Yield: 80%. Yellow solid. M.p = 272–274 ◦ C. Rf = 0.34 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.73 (s, 1H), 11.69 (s, 1H), 11.55 (s, 1H), 7.78–7.54 (m, 4H), 7.02 (d, J = 8.4 Hz, 1H), 6.80 (d, J = 8.3 Hz, 1H), 6.75 (s, 1H), 6.55 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 183.73, 163.04 (d, J = 12.6 Hz), 161.37 (d, J = 12.3 Hz), 161.20, 160.16, 156.65, 155.50, 145.80, 142.25, 137.59, 110.88–110.30 (m), 109.58 (t, J = 25.6 Hz), 107.28, 107.16, 103.82, 99.76, 95.93. 161.41. MS (ESI− ) m/z 417.8 (M − H)− . HRMS (ESI− ): Calcd. for C19 H10 F2 NO6 S− [M − H]− m/z: 418.0202, found: 418.0212. 3-Chloro-N-(1,8-dihydroxy-9-oxo-9H-xanthen-3-yl)-2-fluorobenzenesulfonamide (9h). Yield: 64%. Yellow solid. M.p = 225–227 ◦ C. Rf = 0.31 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.89 (s, 1H), 11.71 (s, 2H), 7.98 (dt, J = 22.0, 7.4 Hz, 2H), 7.68 (t, J = 8.4 Hz, 1H), 7.49 (t, J = 8.1 Hz, 1H), 7.00 (d, J = 8.4 Hz, 1H), 6.79 (d, J = 8.3 Hz, 1H), 6.68 (s, 1H), 6.51 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 183.64, 161.81, 161.26, 160.24, 156.72, 155.58, 153.61 (d, J = 256.5 Hz), 137.57, 136.37, 129.25, 126.13,
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121.68 (d, J = 16.7 Hz), 110.78, 107.30, 107.24, 99.52, 95.62, 95.35, 90.84. MS (ESI− ) m/z 434.0 (M − H)− . HRMS (ESI− ): Calcd. for C19 H14 ClFNO6 S− [M − H]− m/z: 433.9907, found: 433.9917. 2,6-Dichloro-N-(1,8-dihydroxy-9-oxo-9H-xanthen-3-yl)benzenesulfonamide (9i). Yield: 43%. ◦ 1 Yellow solid. M.p = 238–239 C. Rf = 0.37 (Petroleum ether: Acetone = 4:1). H NMR (400 MHz, DMSO-d6 ) δ 11.89 (s, 1H), 11.75 (s, 1H), 11.65 (s, 1H), 7.78–7.53 (m, 4H), 7.02 (d, J = 8.4 Hz, 1H), 6.80 (d, J = 8.1 Hz, 1H), 6.68 (s, 1H), 6.52 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 183.74, 161.26, 160.11, 156.60, 155.49, 145.56, 137.56, 134.78, 134.41, 133.07, 132.15, 110.75, 107.23, 107.19, 103.44, 98.61, 94.86. MS (ESI− ) m/z 450.0 (M−H)− . HRMS (ESI− ): Calcd. for C19 H10 Cl2 NO6 S− [M−H]− m/z: 449.9611, found: 449.9621. N-(1,8-Dihydroxy-9-oxo-9H-xanthen-3-yl)-4-methylbenzenesulfonamide (9j). Yield: 80%. ◦ 1 Yellow solid. M.p = 218–220 C. Rf = 0.35 (Petroleum ether: Acetone = 4:1). H NMR (400 MHz, DMSO) δ 11.71 (s, 2H), 11.33 (brs, 1H), 7.80 (d, J = 8.4 Hz, 2H), 7.70 (t, J = 8.4 Hz, 1H), 7.43 (d, J = 8.0 Hz, 2H), 7.03 (d, J = 8.4 Hz, 1H), 6.80 (d, J = 8.3 Hz, 1H), 6.71 (d, J = 1.9 Hz, 1H), 6.52 (d, J = 1.9 Hz, 1H), 2.35 (s, 3H). 13 C NMR (151 MHz, DMSO) δ 183.44, 161.00, 160.06, 156.48, 155.39, 147.33, 143.78, 137.36, 136.41, 129.88, 126.55, 110.58, 107.07, 107.04, 102.97, 99.33, 95.34, 20.82. MS (ESI− ) m/z 396.0 (M − H)− . HRMS (ESI− ): Calcd. for C20 H14 NO6 S− [M − H]− m/z: 396.0547, found: 396.0553. Yield: 67%. 4-(tert-Butyl)-N-(1,8-dihydroxy-9-oxo-9H-xanthen-3-yl)benzenesulfonamide (9k). ◦ 1 Yellow solid. M.p = 205–208 C. Rf = 0.43 (Petroleum ether: Acetone = 4:1). H NMR (400 MHz, DMSO-d6 ) δ 11.73 (s, 1H), 11.70 (s, 1H), 11.38 (s, 1H), 7.84 (d, J = 8.2 Hz, 2H), 7.73–7.61 (m, 3H), 7.01 (d, J = 8.5 Hz, 1H), 6.79 (d, J = 8.3 Hz, 1H), 6.72 (s, 1H), 6.52 (s, 1H), 1.26 (s, 9H). 13 C NMR (151 MHz, DMSO) δ 183.40, 161.08, 160.13, 156.62, 155.46, 137.36, 136.24, 126.39, 126.36, 110.62, 110.22, 107.12, 106.76, 99.41, 95.35, 95.24, 90.73, 34.83, 30.60. MS (ESI− ) m/z 438.0 (M − H)− . HRMS (ESI− ): Calcd. for C23 H20 NO6 S− [M − H]− m/z: 438.1017, found: 438.1030. 4-Cyclohexyl-N-(1,8-dihydroxy-9-oxo-9H-xanthen-3-yl)benzenesulfonamide (9l). Yield: 66%. Yellow solid. M.p = 203-205 ◦ C. Rf = 0.51 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.71 (s, 1H), 11.69 (s, 1H), 11.36 (s, 1H), 7.83 (d, J = 8.1 Hz, 2H), 7.68 (t, J = 8.4 Hz, 1H), 7.49 (d, J = 8.0 Hz, 2H), 7.01 (d, J = 8.4 Hz, 1H), 6.79 (d, J = 8.3 Hz, 1H), 6.74 (s, 1H), 6.55 (s, 1H), 2.63–2.51 (m, 1H), 1.79–1.60 (m, 5H), 1.44–1.12 (m, 5H). 13 C NMR (151 MHz, DMSO) δ 183.65, 161.15, 160.14, 156.61, 155.47, 153.47, 146.56, 137.52, 136.53, 127.89, 126.72, 110.70, 107.19, 107.15, 103.30, 99.13, 95.19, 43.42, 33.25, 25.99, 25.29. MS (ESI− ) m/z 464.1 (M − H)− . HRMS (ESI− ): Calcd. for C25 H22 NO6 S− [M − H]− m/z: 464.1173, found: 464.1187. N-(1,8-Dihydroxy-9-oxo-9H-xanthen-3-yl)-[1,10 -biphenyl]-4-sulfonamide (9m). Yield: 74%. Yellow solid. M.p = 279–280 ◦ C. Rf = 0.55 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.71 (s, 1H), 11.67 (s, 1H), 11.49 (s, 1H), 8.00 (d, J = 8.3 Hz, 2H), 7.93 (d, J = 8.3 Hz, 2H), 7.74–7.62 (m, 3H), 7.51–7.39 (m, 3H), 7.00 (d, J = 8.3 Hz, 1H), 6.77 (d, J = 6.8 Hz, 2H), 6.59 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 183.59, 161.11, 160.06, 156.54, 155.39, 146.38, 144.86, 137.93, 137.63, 137.45, 128.96, 128.56, 127.68, 127.23, 126.94, 110.63, 107.11, 107.06, 103.34, 99.24, 95.30. MS (ESI− ) m/z 458.0 (M − H)− . HRMS (ESI− ): Calcd. for C25 H16 NO6 S− [M − H]− m/z: 458.0704, found: 458.0718. N-(1,8-Dihydroxy-9-oxo-9H-xanthen-3-yl)naphthalene-1-sulfonamide (9n). Yield: 76%. Yellow solid. M.p = 220–222 ◦ C. Rf = 0.52 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 11.69 (s, 1H), 11.67 (s, 1H), 11.49 (s, 1H), 8.66 (s, 1H), 8.23 (d, J = 7.9 Hz, 1H), 8.15 (d, J = 8.8 Hz, 1H), 8.02 (d, J = 7.9 Hz, 1H), 7.86 (dd, J = 8.7, 1.9 Hz, 1H), 7.78–7.58 (m, 3H), 6.98 (d, J = 8.5 Hz, 1H), 6.82–6.71 (m, 2H), 6.56 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 183.45, 161.02, 160.04, 156.50, 155.38, 147.09, 137.35, 136.20, 134.28, 131.42, 129.77, 129.2-24, 129.13, 128.01, 127.73, 121.56, 110.57, 107.08, 107.04, 103.07, 99.39, 95.44. MS (ESI− ) m/z 432.1 (M − H)− . HRMS (ESI− ): Calcd. for C23 H14 NO6 S− [M − H]− m/z: 432.0547, found: 432.0558. The general synthetic procedures of compounds 15a–15f. The synthetic procedures of 15a–15f were similar to 9n.
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N-(1-Hydroxy-9-oxo-9H-xanthen-3-yl)-[1,10 -biphenyl]-4-sulfonamide (15a). Yield: 60%. Yellow solid. M.p = 234–235 ◦ C. Rf = 0.56 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 12.67 (s, 1H), 11.40 (s, 1H), 8.10 (d, J = 7.6 Hz, 1H), 8.00 (d, J = 8.2 Hz, 2H), 7.93 (d, J = 8.1 Hz, 2H), 7.87 (t, J = 7.8 Hz, 1H), 7.71 (d, J = 7.8 Hz, 2H), 7.63 (d, J = 8.5 Hz, 1H), 7.54 – 7.36 (m, 4H), 6.81 (d, J = 1.8 Hz, 1H), 6.58 (d, J = 1.9 Hz, 1H). 13 C NMR (151 MHz, DMSO) δ 180.18, 161.88, 156.55, 155.41, 145.99, 144.88, 138.02, 137.76, 136.02, 129.03, 128.62, 127.75, 127.28, 127.02, 125.16, 124.57, 119.80, 117.81, 104.30, 99.02, 95.35. MS (ESI− ) m/z 442.1 (M − H)− . HRMS (ESI− ): Calcd. for C25 H16 NO5 S− [M − H]− m/z: 442.0705, found: 442.0716. N-(1-Hydroxy-5-methoxy-9-oxo-9H-xanthen-3-yl)-[1,10 -biphenyl]-4-sulfonamide (15b). Yield: 47%. Yellow solid. M.p = 220–221 ◦ C. Rf = 0.36 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 12.66 (s, 1H), 11.42 (s, 1H), 8.01–7.89 (m, 5H), 7.72 (d, J = 7.7 Hz, 2H), 7.50 (q, J = 8.1, 7.5 Hz, 4H), 7.45–7.34 (m, 1H), 6.80 (s, 1H), 6.58 (s, 1H), 3.97 (s, 3H). 13 C NMR (151 MHz, DMSO) δ 180.21, 161.79, 156.30, 147.88, 145.89, 145.44, 144.87, 138.00, 137.74, 129.02, 128.62, 127.77, 127.21, 127.02, 124.27, 120.56, 116.96, 115.58, 104.28, 99.13, 95.46, 56.21. MS (ESI− ) m/z 471.8 (M − H)− . HRMS (ESI− ): Calcd. for C26 H18 NO6 S− [M − H]− m/z: 472.0860, found: 472.0878. Yield: 48%. N-(1-Hydroxy-5-methyl-9-oxo-9H-xanthen-3-yl)-[1,10 -biphenyl]-4-sulfonamide (15c). Yellow solid. M.p = 235–237 ◦ C. Rf = 0.50 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 12.72 (s, 1H), 11.38 (s, 1H), 8.00 (d, J = 8.4 Hz, 2H), 7.96–7.88 (m, 3H), 7.72 (t, J = 7.9 Hz, 3H), 7.49 (t, J = 7.1 Hz, 2H), 7.43 (d, J = 6.6 Hz, 1H), 7.36 (t, J = 7.7 Hz, 1H), 6.83 (s, 1H), 6.59 (s, 1H), 2.48 (s, 3H). 13 C NMR (151 MHz, DMSO) δ 180.38, 161.75, 156.36, 153.68, 145.88, 144.79, 137.95, 137.74, 136.61, 128.97, 128.56, 127.67, 127.19, 126.95, 126.74, 123.93, 122.68, 119.55, 104.05, 99.03, 95.57, 15.06. MS (ESI− ) m/z 456.1 (M − H)− . HRMS (ESI− ): Calcd. for C26 H18 NO5 S− [M − H]− m/z: 456.0911, found: 456.0926. N-(1-Hydroxy-7-methoxy-9-oxo-9H-xanthen-3-yl)-[1,10 -biphenyl]-4-sulfonamide (15d). Yield: 58%. Yellow solid. M.p = 220–222 ◦ C. Rf = 0.33 (Petroleum ether: Acetone = 4:1). 1H NMR (400 MHz, DMSO-d6) δ 12.62 (s, 1H), 11.28 (s, 1H), 7.95–7.77 (m, 4H), 7.70–7.55 (m, 2H), 7.45–7.26 (m, 6H), 6.71 (s, 1H), 6.48 (s, 1H), 3.77 (s, 3H). 13C NMR (151 MHz, DMSO) δ 179.82, 161.68, 156.45, 155.70, 150.07, 145.72, 144.81, 137.95, 137.70, 128.96, 128.56, 127.68, 127.20, 126.95, 124.93, 120.11, 119.34, 105.03, 103.91, 98.80, 95.13, 55.63. MS (ESI− ) m/z 471.9 (M − H)− . HRMS (ESI− ): Calcd. for C26H18NO6S− [M − H]− m/z: 472.0860, found: 472.0873. N-(1-Hydroxy-7-methyl-9-oxo-9H-xanthen-3-yl)-[1,10 -biphenyl]-4-sulfonamide (15e). Yield: 60%. Yellow solid. M.p = 239–240. Rf = 0.48 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 12.72 (s, 1H), 11.39 (s, 1H), 7.99 (d, J = 8.4 Hz, 2H), 7.93 (d, J = 8.5 Hz, 2H), 7.87 (s, 1H), 7.71 (d, J = 7.2 Hz, 2H), 7.66 (d, J = 8.7 Hz, 1H), 7.56–7.33 (m, 4H), 6.79 (d, J = 1.8 Hz, 1H), 6.56 (d, J = 1.7 Hz, 1H), 2.40 (s, 3H). 13 C NMR (151 MHz, DMSO) δ 180.10, 161.81, 156.47, 153.59, 145.80, 144.80, 137.95, 137.71, 137.01, 134.02, 128.96, 128.55, 127.68, 127.21, 126.95, 124.29, 119.36, 117.55, 104.20, 98.84, 95.22, 20.12. MS (ESI− ) m/z 456.1 (M − H)− . HRMS (ESI− ): Calcd. for C26 H18 NO5 S− [M − H]− m/z: 456.0911, found: 456.0928. N-(7-Chloro-1-hydroxy-9-oxo-9H-xanthen-3-yl)-[1,10 -biphenyl]-4-sulfonamide (15f). Yield: 58%. Yellow solid. M.p = 259–260 ◦ C. Rf = 0.38 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 12.42 (s, 1H), 11.45 (s, 1H), 8.05–7.95 (m, 3H), 7.96–7.86 (m, 3H), 7.75–7.66 (m, 3H), 7.49 (t, J = 7.3 Hz, 2H), 7.43 (d, J = 7.4 Hz, 1H), 6.81 (d, J = 2.0 Hz, 1H), 6.59 (d, J = 2.0 Hz, 1H). MS (ESI− ) m/z 476.0 (M − H)− . HRMS (ESI− ): Calcd. for C25 H15 ClNO5 S− [M − H]− m/z: 476.0365, found: 476.0379. N-(7-Fluoro-1-hydroxy-9-oxo-9H-xanthen-3-yl)-[1,10 -biphenyl]-4-sulfonamide (15g). Yield: 51%. ◦ 1 Yellow solid. M.p = 250–252 C. Rf = 0.45 (Petroleum ether: Acetone = 4:1). H NMR (400 MHz, DMSO-d6 ) δ 12.46 (s, 1H), 11.45 (s, 1H), 7.99 (d, J = 8.4 Hz, 2H), 7.93 (d, J = 8.8 Hz, 2H), 7.82–7.68 (m, 5H), 7.56–7.36 (m, 3H), 6.81 (s, 1H), 6.58 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 179.36, 161.61, 158.10 (d, J = 243.9 Hz), 156.53, 151.81, 146.17, 144.83, 137.93, 137.67, 128.97, 128.57, 127.69, 127.21, 126.96, 123.81 (d, J = 24.7 Hz), 120.76 (d, J = 7.4 Hz), 120.40 (d, J = 7.9 Hz), 109.78 (d, J = 24.3 Hz), 103.86,
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99.07, 95.22. MS (ESI− ) m/z 460.1 (M − H)− . HRMS (ESI− ): Calcd. for C25 H15 FNO5 S− [M − H]− m/z: 460.0660, found: 460.0677. N-(1-Hydroxy-7-nitro-9-oxo-9H-xanthen-3-yl)-[1,10 -biphenyl]-4-sulfonamide (15h). Yield: 35%. Yellow solid. M.p = 243-245. Rf = 0.25 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 12.21 (s, 1H), 11.51 (s, 1H), 8.76 (s, 1H), 8.64–8.55 (m, 1H), 8.00 (d, J = 8.1 Hz, 2H), 7.93 (d, J = 8.3 Hz, 2H), 7.86 (d, J = 9.6 Hz, 1H), 7.71 (d, J = 7.7 Hz, 2H), 7.48 (t, J = 7.6 Hz, 2H), 7.47–7.38 (m, 1H), 6.85 (s, 1H), 6.64 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 178.84, 161.67, 158.60, 156.39, 146.65, 144.95, 143.43, 137.97, 137.65, 129.79, 129.03, 128.64, 127.77, 127.28, 127.01, 121.11, 120.15, 119.90, 104.27, 99.76, 95.72. MS (ESI− ) m/z 487.1 (M − H)− . HRMS (ESI− ): Calcd. for C25 H15 N2 O7 S− [M − H]− m/z: 487.0605, found: 487.0626. N-(1,7-Dihydroxy-9-oxo-9H-xanthen-3-yl)-[1,10 -biphenyl]-4-sulfonamide (15i). To a solution of Compound 15d (30 mg, 0.063 mmol) in dry DCM (5 mL), BBr3 (1 mol/L in DCM, 0.5 mL) was added. The reaction was kept at 0 ◦ C for 2 h and then quenched with methanol (10 mL). The mixed solution was concentrated in vacuo and the residue was purified by flash column chromatography to give the demethylated product 15i as a yellow solid (18 mg, yield: 61%). Yellow solid. M.p = 257–259 ◦ C. Rf = 0.53 (Petroleum ether: Acetone = 2:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 12.74 (s, 1H), 11.34 (s, 1H), 10.05 (s, 1H), 7.98 (d, J = 9.5 Hz, 2H), 7.92 (d, J = 9.3 Hz, 2H), 7.71 (d, J = 8.1 Hz, 2H), 7.56–7.13 (m, 6H), 6.77 (s, 1H), 6.54 (s, 1H). 13 C NMR (151 MHz, DMSO) δ 179.99, 161.71, 156.49, 153.97, 148.98, 145.60, 144.79, 137.96, 137.71, 128.96, 128.55, 127.67, 127.19, 126.96, 124.87, 120.28, 119.06, 107.69, 103.86, 98.65, 95.08. MS (ESI− ) m/z 458.1 (M − H)− . HRMS (ESI− ): Calcd. for C25 H16 NO6 S− [M − H]− m/z: 458.0704, found: 458.0710. 6-([1,10 -Biphenyl]-4-sulfonamido)-8-hydroxy-9-oxo-9H-xanthen-2-yl acetate (15j). To a solution of Compound 15i (19 mg, 0.041 mmol) and NEt3 (0.017 mL, 0.124 mmol) in dry DCM (5 mL), Ac2 O (0.006 mL, 0.062 mmol) was added. After the reaction was complete, which was monitored by TLC, the reaction mixture was concentrated in vacuo and the residue was purified by flash column chromatography to give the acetylated product 15j as a yellow solid (16 mg, yield: 77%). Yellow solid. M.p = 261–262 ◦ C. Rf = 0.51 (Petroleum ether: Acetone = 4:1). 1 H NMR (400 MHz, DMSO-d6 ) δ 12.52 (s, 1H), 11.45 (s, 1H), 8.00 (d, J = 8.2 Hz, 2H), 7.93 (d, J = 8.3 Hz, 2H), 7.82 (s, 1H), 7.77–7.61 (m, 4H), 7.53–7.37 (m, 3H), 6.81 (s, 1H), 6.58 (s, 1H), 2.30 (s, 3H). 13 C NMR (151 MHz, DMSO) δ 179.65, 169.13, 161.74, 156.59, 152.90, 146.54, 146.06, 144.90, 138.00, 137.70, 130.13, 129.03, 128.63, 127.77, 127.27, 127.02, 120.30, 119.31, 117.22, 104.04, 99.05, 95.30, 20.73. MS (ESI− ) m/z 500.0 (M − H)− . HRMS (ESI− ): Calcd. for C27 H18 NO7 S− [M − H]− m/z: 500.0809, found: 500.0831. 3.2. Biological Evaluation Assay 3.2.1. In Vitro PGAM1 Enzyme Inhibitory Activity Assay First, 1 µL of test compound (dissolved in DMSO) with a specific concentration, 1 µL of recombinant PGAM1 and 48 µL of Tris buffer solution (50 mM, pH 8.0), was added to a 96-well plate followed by preincubation for 2 min at room temperature. Then, 49 µL of enzyme buffer (containing 0.5 U/mL enolase, 0.5 U/mL PK, 0.1 U/mL LDH, 5 mM MgCl2 , 1 mM ADP, 100 mM KCl, 0.2 mM NADH, 100 mM Tris pH 8.0) was added to the mixture. Finally, 1 µL of 3PG solution (200 mM) was added to initiate the reaction. The decrease in OD (λ = 340 nm) from the oxidation of NADH was measured by a microplate reader as PGAM1 activity. 3.2.2. Counter-Screen Assay Activities of the Selected Compounds First, 1 µL of test compound (dissolved in DMSO) with a specific concentration and 49 µL of Tris buffer solution (50 mM, pH 8.0) were added to a 96-well plate followed by preincubation for 2 min at room temperature. Then, 49 µL of enzymes buffer (containing 0.5 U/mL enolase, 0.5 U/mL PK, 0.1 U/mL LDH, 5 mM MgCl2 , 1 mM ADP, 100 mM KCl, 0.2 mM NADH, 100 mM Tris pH 8.0)
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was added to the mixture. Finally, 1 µL of 2PG solution (200 mM) was added to initiate the reaction. The decrease in OD (λ = 340 nm) from the oxidation of NADH was measured as counter-screen assay activities by a microplate reader. 3.2.3. In Vitro Antiproliferation of H1299 Cell Activity Assay 2 × 103 cells were seeded in a 96-well plate before starting the assay and they were cultured at 37 ◦ C. After seeding for 24 h, cells were treated with inhibitors with a specific concentration and incubated at 37 ◦ C for 72 h, followed by incubation with 0.5 mg mL−1 MTT for 4 h at 37 ◦ C. Then, 200 µL of DMSO was added and the absorbance was measured at 570 nm. 4. Conclusions In summary, based on our previous work, we continued two rounds of modification to discover new N-xanthone benzenesulfonamides as PGAM1 inhibitors. A total number of 24 N-xanthone benzenesulfonamides were designed and synthesized, and their inhibitory activities against PGAM1 were evaluated. Among them, the most active and specific compound 15h (IC50 = 2.1 µM) showed a 5-fold enhancement of PGAM1 inhibitory activity and a much higher specificity compared to PGMI-004A. Further, the antiproliferation activities on the H1299 cell line of the representative N-xanthone benzenesulfonamides were also evaluated, which showed a slightly increased antiproliferative activity. Consequently, in this study, we have expanded the structural types of PGAM1 inhibitors and provided a new direction for further development of more efficient PGAM1 inhibitors. Author Contributions: Conceptualization, P.W., D.Y., and L.Z.; Chemistry, P.W.; Biological experiments, L.J.; Molecular Docking, Y.C.; Writing, Review, and Editing, P.W., L.Z., and D.Y. Funding: This research was funded by the Chinese National Natural Science Foundation (No. 21472026), the Shanghai Municipal Committee of Science and Technology (No. 14XD1400300) and the program for Shanghai Rising Star (No. 15QA1400300). Conflicts of Interest: The authors declare no conflict of interest.
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Sample Availability: Samples of the compounds are not available from the authors. © 2018 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
