Lewis Base-Catalyzed Intermolecular Triazene Alkyne Cycloaddition for Late-Stage Functionalization and Scaffold Diversiﬁcation

ARTICLE

https://doi.org/10.1038/s42004-019-0168-6 OPEN Lewis base-catalyzed intermolecular triazene alkyne cycloaddition for late-stage functionalization and scaffold diversiﬁcation

Shuaipeng Lv1,4, Hui Zhou2,4, Xin Yu1,2, Yue Xu1, Huijuan Zhu1, Min Wang1, Haitao Liu1, Ziru Dai1, Guibo Sun1,

1234567890():,; Xiaojie Gong3, Xiaobo Sun1 & Lei Wang1

3-Trifluoromethylpyrazole and its derivatives are of major interest to both the agrochemical and pharmaceutical industry for their diverse biological activities. Reported routes for the synthesis of 3-trifluoromethylpyrazoles are hindered by poor regioselectivity and limited scope of application. Here we report a directed Lewis base catalyzed intermolecular triazene- alkyne cycloaddition. It is featured that the combination of 1,8-diazabicyclo[5.4.0]undec-7- ene and 2,2,2-trifluorodiazoethane produces reactive triazene intermediates, which readily participate in cycloaddition reactions with terminal/internal alkynes, thus assembling densely substituted 3-trifluoromethylpyrazole scaffolds with environmental friendliness and operational simplicity. Synthetic utility of the protocol is highlighted by late-stage functionalization and scaffolds diversification. The practical value is also emphasized in potential platelet aggregation inhibitor synthesis.

1 Beijing Key Laboratory of Innovative Drug Discovery of Traditional Chinese Medicine and Translational Medicine, Institute of Medicinal Plant Development, Chinese Academy of Medical Science & Peking Union Medical College, 100193 Beijing, China. 2 Department of Chemistry and Pharmacy, Zhuhai College of Jilin University, 519041 Zhuhai, China. 3 College of Life Science, Dalian Minzu University, 116600 Dalian, China. 4These authors contributed equally: Shuaipeng Lv, Hui Zhou. Correspondence and requests for materials should be addressed to X.S. (email: [email protected])ortoL.W.(email:[email protected])

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yrazoles are the core scaffolds of numerous biologically the use of stoichiometric catalyst or prefunctionalized starting Pactive molecules and exhibit innumerable applications in materials and the explored methods are confined to a narrow range chemistry and biology. Pyrazole derivatives represent one of of substrates and limited scope of general applications. Due to the 34 the most active classes of compounds and possess a broad range explored N-terminal electrophilicity of CF3CHN2 and triazenes as of chemical, biological, agrochemical, and pharmacological a versatile tool in organic synthesis41 together with the convenient 1,2 fl properties . 3-Tri uoromethylpyrazoles, well-known examples methods for the synthesis of CF3CHN2 in different solvents of pyrazole derivatives, are key privileged scaffolds widely existed reported by Ma’sgroup42, the applications of the merger of N- in many important biologically active molecules, agrochemicals, terminal electrophilicity of CF3CHN2 and Lewis base to form and pharmaceuticals (Fig. 1a)3–11. Conventional approaches for triazene intermediates in organic synthesis and medicinal chemistry the construction of 3-trifluoromethylpyrazoles involve by the have rarely been studied. condensation of hydrazines with fluoroalkyl 1, 3-dicarbonyl Late-stage functionalization (LSF)43–46, a valuable tool for compounds (Fig. 1b)12–15. However, these methods are limited by directly introducing functional groups onto a bioactive compound, the need for prefunctionalized starting materials and by poor has merged as an important strategy for contemporary drug dis- regioselectivities. Notwithstanding recent progress, these specific covery due to enabling rapid structural diversity of drug candidates methods are incompatible with the extreme value of 3- or drug-like molecules to ultimately affect their physiochemical trifluoromethylpyrazoles. properties such as ADME (absorption, distribution, metabolism, 47 Recently, 2,2,2-trifluorodiazoethane (CF3CHN2) has emerged as and excretion) . In support of an on-going drug discovery strategy, an attractive synthon and has been extensively studied as a metal novel LSF with high efficiency, selectivity, and operational simpli- carbene precursor, 1,3-dipole, C- nucleophile/electrophile, and N- city is highly desirable. 3-Trifluoromethylpyrazoles, as COXs inhi- terminal electrophile16–40. Compelling examples have been reported bition pharmacophore8–10,48, their derivatives have been screened fl on the utilization of CF3CHN2 as a uorine-containing building as clinical drug candidates and commercial pharmaceuticals. Drugs block. Considerable efforts have also been expended on the con- aimed at COXs inhibition is a billion opportunity, which has been struction of 3-trifluoromethylpyrazoles25,30. This has been chal- inspiring medicinal chemists to search constantly for novel COX lenging because the effective documented processes always rely on inhibitors. The attractive transition-metal-free transformation from

a F3C

F C R3 Major utilities 3 N Celecoxib N Agrochemicals Me N R2 COX-2 inhibitor pharmaceuticals N R1

SO2NH2

b O O Cyclocondensation Ag2O or Base R2 H 2 F3C R 1 R –NHNH2 CF3CHN2 3 4 F3C R /CH2R /H Scope Stoichiometric catalyst regioselectivity N 2 substrate dependent 3 N R /EWG/H R EWG EWG NNHR1 R1 2 Togni reagent R CF3CHN2 R4 NO2 H Ag2O or Base R3

c Hypothesis + DBU N Activated CF CHN + N 3 2 N CF3 N DBU Triazene

Work overview F C 2 + DBU 3 R Lewis base catalysis Transition-metal free 1 2 N R R N CF 3 N R1 Terminal/ N H Internal alkyne Triazene

CF3 >70 Examples, yield up to 98% Late-stage Drug Broad substrates scope fuctionalization X N candidates Simple & mild reaction conditions Scaffolds n N development diversification H Functional gropus and scaff olds diversity Pharmaceutical application X = C,O, N; n (<10) = H/functional group = Pharmaceuticals; natural products; bioactive molecules

Fig. 1 Recent directions and envisaged approach for 3-trifluoromethylpyrazoles synthesis. a Importance of 3-trifluoromethylpyrazoles. b Known protocols of 3-trifluoromethylpyrazoles synthesis and limitations. c Proposed triazene-alkyne cycloaddition for the direct [3 + 2] assembly of 3- trifluoromethylpyrazoles, late-stage functionalization, and scaffolds diversification

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Table 1 Selected optimization of reaction conditions

Entry Variations from the ‘initial conditions' Yield (%)a 1 No DBU <10 2 1,1,3,3-tetramethylguanidine 57 instead of DBU 3 Triethylamine instead of DBU 65 4 4-dimethylaminopyridine instead of DBU 35 5 Tetramethylethylenediamine 51 instead of DBU 6 1,4-diazabicyclo[2.2.2]octane 48 instead of DBU 7CF3CHN2 in 1,4-dioxane 69 8CF3CHN2 in DCE 22 9 MeCN instead of 1,4-dioxane 14 10 Diethyl ether instead of 1,4-dioxane 31 11 70 °C 79 12 80 °C 97 13 10 mol% DBU, 80 °C 72

Unless otherwise speciﬁed, all reactions were carried out using phenylacetylene 1a (0.3 mmol, 1.0 equiv.) and CF3CHN2 2 (1.2 mmol, 4.0 equiv., 1.5 M in solvents), 20 mol% base (0.06 mmol, 0.2 equiv.), solvent (0.4 mL), 12 h aYield of the isolated product 3a after chromatography

the alkyne moiety embedded compounds with known biological external solvent (71%) (see Supplementary Table 1). Various properties to the corresponding 3-trifluoromethylpyrazole deriva- bases were then evaluated, revealing that DBU was still essential tives via LSF and scaffolds diversification is still rarely explored. for the high efficiency of this transformation (Table 1, entry 2–6). We hypothesized that by the combination of Lewis base cata- Further optimizations of reaction conditions indicated that the fi lysis and CF3CHN2 to generate reactive triazene intermediates stock solvents of CF3CHN2 could signi cantly affect the reaction could be employed in cycloaddition reactions with terminal/ with decreased yield (Table 1, entry 7, 8). Attempts to increase the internal alkynes and open a new avenue for the assembly of yield of 3a were carried out with different reaction temperatures. densely functionalized 3-trifluoromethylpyrazoles. Also, the To our delight, this transformation gave the almost quantitative newly developed transformations could be hypothetically yield at 80 °C (Table 1, entry 12). It was noted that the erosive expanded to enable LSF of pharmaceuticals and diversification of yield was observed when the catalyst loading was reduced to 10 clinical drugs, natural products, and bioactive molecules with 3- mol% (Table 1, entry 13). trifluoromethylpyrazole scaffold. Currently, there are more than 400 drugs and more than 4000 natural products bearing alkyne fl moiety (http://dnp.chemnetbase.com). Thus, there is a huge Synthesis of 3-tri uormethylpyrazoles. With the established optimal reaction conditions in hand, the generality of this potential demand to develop a facile and generally applicable 3- fl trifluoromethylpyrazole synthetic procedure for LSF and scaffolds approach to synthesize a range of 3-tri uormethylpyrazoles was fi evaluated (Fig. 2). Various terminal alkynes 1 bearing electron- diversi cation. fi Here we show a transition-metal free catalytic intermolecular neutral, electron-rich, and electron-de cient substituents on the aromatic ring were found to be suitable for this reaction to form triazene-alkyne cycloaddition (TAC) procedure for the synthesis – of highly substituted 3-trifluoromethylpyrazoles and efficient the corresponding pyrazoles (3a q) with very good to excellent installation of the title heterocycle into complex bioactive mole- yields. Notably, heteroaryl terminal alkynes, electron-poor alkyne, cules in the context of LSF and scaffold diversification. Con- and N-protected aliphatic alkyne were also readily converted into fi 3r–x sidering the important role of COXs inhibition in antiplatelet the desired products with high ef ciency ( ). However, low therapy, we also extend the protocol to assemble drug-like platelet yields were obtained when aliphatic alkynes were used in this 3y 3z aggregation inhibitors (Fig. 1c). transformation ( , ). From the perspective of product diversity, internal alkynes 4 were also explored for this DBU-catalyzed TAC strategy (Fig. 2). Results The alkyne substrates bearing phenyl, ester, phosphonate diester, Screening of reaction conditions. The designed Lewis base cat- aldehyde, halides, indole, N, N-dimethylacetamide, N, N- alyzed TAC reaction was evaluated by using the simple terminal dimethylethanethioamide, 2-pyridyl, methyl, and trifluoromethyl alkyne 1a (1.0 equiv.) and CF3CHN2 2a (4.0 equiv., 1.5 mol/L in groups were all compatible, resulting in the synthesis of various toluene) in the presence of DBU as Lewis bases at 60 °C. We desired densely functionalized 3-trifluormethylpyrazoles with found that the reaction proceeded smoothly and the cycloadduct moderate to excellent yields (5a–m). However, erosive yields 3a was obtained in good yield when 1,4-dioxane was used as the were obtained with diphenylacetylene and phenyl

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a F C R2 R2 DBU 20 mol% 3 1,4-dioxane, 80°C + CF3CHN2 N R1 R1 in toluene N H 1: Terminal alkynes 2 3/5 4: Internal alkynes b F C F C F C 3 3 R 3 F3C F C Me 3 3b, R = Me, 84%, 12 h 3i, R = F, 87%, 12 h N N N 3c, R = OMe, 92%, 15 h N N N 3j, R =CI, 92%, 12 h N N H H N H 3d, R = t-Bu, 93%, 12 h 3k, R =Br, 90%, 12 h N H R H R 3e, R = CF , 98%, 14 h 3g, R = Me, 89%, 12 h 3l, R =NO , 75%, 12 h 3a, 97%, 12 h 3 3f, 96%, 12 h 2 3h, R = OMe, 87%, 12 h

F C F C F C 3 3 F C F3C 3 F C F C 3 CI 3 Br 3 Me N N N N N N N OMe N N N N N N N H H CI H N N H H H H

3m, 97%, 12 h 3n, 95%, 12 h 3o, 95%, 12 h 3p, 79%, 18 h 3q, 94%, 29 h 3r, 96%,16 h 3s, 94%,12 h

F C F C 3 3 F C F C F C 3 3 F3C 3 F3C H N N N N N Ph N N N N N n-C H N CO Et N N 5 11 N H S H 2 S H NH N H H H H 3z, 26%,36 h 3t, 79%, 15 h 3u, 86%, 15 h 3v, 78%, 12 h 3w, 83%, 32 h 3x, 71%, 15 h 3y, 31%, 24 h

c CO Me 2 X Me O F C R OEt 3 F C N F C CO Et F C CHO 3 3 2 F3C P 3 OEt Me N F C N Ph 3 N N N N N Ph Ph Ph Ph H N N N N H H N H H Ph 5d, R = CI, 64%, 12 h N 5h, X = O, 60%, 48 h 5e, R = Br, 65%, 12 h H 5i, X = S, 57%, 48 h 5a, 82%, 20 h 5b, 79%, 24 h 5c, 52%, 12 h 5f, R = I, 51%, 12 h 5g, 56%, 24 h

F C CO Et F C SePh Ph 3 2 3 F3C F3C Me F C CF3 3 F3C CO2Me

N N N N N CO Me N N N Ph N Ph N 2 N CO2Me CO Me H N N 2 H H H H H 5j, 71%, 20 h 5k, 58%, 24 h 5I, 65%, 12 h 5m, 93%,12 h 5n, 35%,48 h 5o, 31%,48 h

Fig. 2 Scope of terminal and internal alkynes. a Reaction scheme. b Terminal alkynes. c Internal alkynes. Reaction conditions: alkynes 1 or 4 (0.3 mmol, 1.0 equiv.) and CF3CHN2 2 (1.2 mmol, 4 equiv., 1.5 M in toluene), DBU (0.06 mmol, 20 mol%), 1,4-dioxane (0.4 mL), 80 °C, 12–32 h; yields of isolated products 3 or 5 after chromatography

(phenylethynyl) selane as substrates (5n, 5o). Notably, all the 3-trifluormethylpyrazole into various kinds of bioactive internal alkynes afforded densely functionalized 3- compounds, ranging from pharmaceutically relevant molecules trifluormethylpyrazoles with high levels of regioselectivity. Based and natural products to a panel of bioactive heterocycles. As on the X-ray crystallographic analysis of compounds 5a and 5k, indicated in Fig. 4, paciltaxel, hydroxycamptothecin, and the configuration of the functionalized 3-trifluormethylpyrazoles fluorouracil, used to treat different types of cancer, was products was assigned (see Supplementary Figs. 230 and 231). directly diversified by TAC strategy in very good yield (9a–d). However, 5k and 5l possessed distinctive substituents compared Similar method was also directly applied to diversify efavirenz with others functionalized pyrazoles mainly due to electronic and and yielded the product with high efficiency and mono/di steric reasons. selectivity (9e, 9f). Interestingly, penicillin G, a commercially available antibiotic used to treat bacterial infections, was also Late-stage functionalization. To further indicate the utility of functionally obtained in good yield. Artemisinin, medication TAC procedures, we set out to perform LSF of pharmaceutically approved for the treatment of malaria, could be varied in a TAC relevant molecules. As indicated in Fig. 3, erlotinib, used to treat manner and was readily reached in 45% yield after this diver- fi nonsmall cell lung cancer and pancreatic cancer, was directly si cation (9h). fi functionalized by our TAC strategy in 49% yield (7a). Further- The scaffold diversi cation strategy in terms of various more, efavirenz, a commercially available anti-HIV drug, was also natural products was also explored. As shown in Fig. 4, natural functionally obtained with product diversity in good yield (7b). In products related with tetracyclic and pentacyclic triterpenes addition, the extension of the LSF to pargyline, an anti- such as cholesterol and oleanolic acid were tolerated for this fi hypertensive drug, was also achieved and formed the corre- diversi cation with high yields (9i, 9j). The variation of scaffold fl sponding derivative with good yield (7c). range to avonoid and coumarin was also successful and assembledtherelateddiversified products with very good yields (9k, 9l). Lignins and alkaloids, such as podophyllotoxin Scaffold diversification. To show the generality of TAC pro- and huperzine A, were also investigated in this TAC cedure, we set out to perform scaffold diversification to embed

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Standard CF3 CF3CHN2 coditions + in toluene Late-stage N fuctionalization N 62 H 7 b F3C H N Cl N F C 3 Me N N F3C N N H F3C N N N H N O NH H O O O O 7a, 49%, 24 h O 7b, 52%, 30 h 7c, 59%, 24 h Me from erlotinib from efavirenz from pargyline Me

Fig. 3 Late-stage functionalization. a Reaction scheme. b Late-stage functionalization of marketed drugs. Reaction conditions: 6 (0.3 mmol, 1.0 equiv.) and

CF3CHN2 2 (1.2 mmol, 4 equiv., 1.5 M in toluene), DBU (0.06 mmol, 20 mol%), 1,4-dioxane (0.4 mL), 80 °C, 24-30 h; yields of isolated products 7 after chromatography

a Standard CF3 conditions x CF CHN X + 3 2 N n in toluene Scaffolds n N diversification H 8 2 9 X = C,O, N; n (1–4) b Anti-cancer analogues Anti-HIV analogues F C O O F3C 3 AcO F F Cl O HN N CF3 N OAc N F C O 3 O F C CF3 O F C 3 HN N O N NH 3 O H N O N O N F3C 4 O N N N H N N N NH F3C F C N Cl H N H O O 3 H O OH O O N NH N NH O O Ph OH O O [a] 9e, 51% 15 h[b] 9f, 43% 15 h[b] 9a, 70%, 24 h 9b, 79%, 24 h 9c, 71%, 24 h 9d, 75%, 24 h from fluorouracil from efavirenz from efavirenz from paclitaxel from hydroxycamptothecin from fluorouracil

Antibiotic and antimalarial analogues c Triterpenes Flavonoid Courmarin

H H F3C N S O N O O NH N O F3C O O F C F3C 3 N H CO Bn O O N NH 2 O O H O N NH O H O N H H H O CF3 F C NH H 3 N O HN N O O 9g, 55%, 24 h 9h, 45%, 18 h 9i, 70%, 24 h 9j, 63%, 20 h 9k, 85%, 18 h 9l, 75%, 12 h from penicillin from artemisinin from cholestrol from oleanolic acid from flavonoid from courmarin

Lignin Alkaloids d Bioactive heterocycles

O O OMe O F C H 3 HH F3C NH CF3 F3C X OMe N N O O N N N HN N N N O H N N OMe N H H H 9q, 90%, 24 h 9r, 96%, 24 h O O F C NH OMe 3 N from indole from carbazole 9m, X = NH, 41%, 24 h 9n, X = O,53%, 12 h 9o, 61%, 24 h 9p, 78%, 24 h from podophyllotoxin from huperzine A from tetrahydropalmatine NH2 O O O N N O O F3C O F3C O F C N N F3C 3 N N N N N N N N N H N H NH N N F C NH O H 3 N H N F3C 9s, 86%, 24 h 9t, 79%, 24 h 9u, 91%, 24 h 9v, 85%, 24 h 9W, 80%, 24 h 9x, 73%, 18 h from dihydroquinolinone from acridone from azepine from dihydroazepine from adenine from glucofuranose

Fig. 4 Scaffold diversification. a Reaction scheme. b Diversification of related drug analogs. c Diversification of related natural products. d Diversification of bioactive heterocycles. Reaction conditions: 8 (0.3 mmol, 1.0 equiv.) and CF3CHN2 2 (1.2 mmol, 4 equiv., 1.5 M in toluene), DBU (0.06 mmol, 20 mol%), a b 1,4-dioxane (0.4 mL), 80 °C, 24–30 h; yields of isolated products 9 after chromatography. CF3CHN2 2 (2.4 mmol, 8 equiv., 1.5 M in toluene). CF3CHN2 2 (2.4 mmol, 8 equiv., 1.5 M in toluene), and mono/di selectivity (9e, 9f)

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F3C 3b: R = Me, 5 mmol, 0.965 g, 85% DBU 20 mol% 3c: R = OMe, 5 mmol, 1.125 g, 93% CF3CHN2 1,4-dioxane, 80 °C, 12 h + in toluene N 3i: R = F, 5 mmol, 1.012 g, 88% N 3j: R = CI, 5 mmol, 1.131 g, 92% H R >10 g scale, 3c, 60 mmol, 13.07 g, 92%, 60 h R

b F3C

F3C N N N R Ref. [48] N CI Ref. [49] 3j 3b or 3i R = F, SC-560 Mavacoxib OOS COX-1 inhibitor R = Me, OMe NH2 Celecoxib

Fig. 5 Synthetic utility of TAC strategy. a Gram-scale and 10-g-scale synthesis. b Pharmaceuticals synthesis

a F3CF3C F C F C MeO O 3 3 CI Drug candidate COX-I/II N N N N inhibition N NH development R N N N CI OMe S TAC application N R1 N N N

P2Y12-receptor Clopidogrel S S S S antagonism 10a 10b 10c

b F3C F3C N N NH DBU 10 mol% N N 2 1,4-dioxane, 80 °C Ref. 48 N R + N H CF N S 3 S in toluene S 10b, R = CI, 64%, 36 h ′ 1a 10a, 65%, 24 h 10c, R = OMe, 62%, 30 h c

Val 349 Val 349 Tyr 385 Ser 530 2.4 Å 2.6 Å

2.6 Å Val 349 2.8 Å 2.2 Å Tyr 385 Ser 530 Tyr 385 2.0 Å

Ser 530

Compound 10a Compound 10b Compound 10c

d 100

20 Platelet aggregation (%)

0 124 124 124 124 (mM)

Arachidonic acid 10a 10b 10c Aspirin

Fig. 6 Application of TAC strategy for drug-like molecule development. a Envisaged TAC method for the synthesis of antiplatelet drug-like molecules. b Synthesis of target molecules 10a–c. c Stereo diagram of binding conformation of novel analogs 10a–c with COX-1 isozyme (PDB:3N8Y). d The Platelet aggregation inhibition assay (error bars means ± standard deviation from three independent experiments)

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a N F3C D N F3C D N 2 Standard N Ph N + conditions H 3a H CF3 N Ph N N2 in toluene 1,3-H Ph H 1a’ 2 3a’ H CF3 shift in toluene F C H N-electrophilicity 3 2 N N Ph N N III + + N 1a N N N 2 rt, 12 h N CF + DBU + F C N 3 CF N 3 N H 3 I 0.06 mmol N in CDCI3 N N “Triazene 0.8 ml F3C intermediate” Ph IV II 7.59 7.59 7.57 7.50 7.50 7.50 7.49 7.49 7.48 7.48 7.45 7.45 7.45 7.44 7.26 6.80

10.73 b F3C D/H 90/10

N N Ph H 3a’ 1.08 1.93 1.90 1.00 0.10

10.5 10.0 9.5 9.0 8.5 8.0 7.5 7.0 6.5 6.0 5.5 5.04.5 4.0 3.5 3.0 2.5 2.0 1.51.0 0.5 0.0 1.94 1.94 1.93 1.72 1.65 4.44 4.44 4.43 4.45 3.39 3.37 3.36 3.35 3.34 3.33 2.71

7.26 c

N + N N F3C N 2.00 5.91 1.92 2.08 4.39 2.33

7.0 6.5 6.0 5.5 5.04.5 4.0 3.5 3.0 2.5 2.0 1.51.0 0.5 0.0

Fig. 7 Postulated mechanism for the TAC reactions. a Plausible catalytic cycle. b NMR study of deuterium-labeling experiment. c NMR study of DBU-

CF3CHN2 interaction

transformation, giving the corresponding functionalized deri- nucleotides found in both DNA and RNA, was also suitable for vatives with moderate to good yields (9m–o). Interestingly, this diversification and afforded the functional derivative in very tetrahydropalmatine, applications as an adrenergic agent and a good yield (9w). Moreover, we also extended the strategy to dopaminergic antagonist, was additionally found to be suitable protected glucofuranose and provided the diversified product forthisdiversification in 78% yield (9p). with high efficiency (9x). Beyond scaffolds diversification of highly valuable pharmaceuticals and natural products. This TAC strategy also unlocked new pathways for the straightforward and efficient diversification of Applications. Gram-scale synthesis of 3b, 3c, 3i, and 3j using the appealing bioactive molecules. Privileged N-containing hetero- established TAC system promoted smoothly without affecting the cycles with various biological activities, such as indole, carbazole, efficiency outcome of the reactions. To further highlight the dihydroquinolinone, acridone, azepine, and dihydroazepine were potential industrial application of this transformation, a 10-g- also engaged in this diversification to construct the corresponding scale synthesis of 3c proceeded smoothly without erosion of the functionalized products in high to excellent yields (9q–v). yield but only with increased reaction time (Fig. 5). In addition, Furthermore, adenine, a fundamental component of adenine the gram-scale products, as crucial intermediates, can be further

COMMUNICATIONS CHEMISTRY | (2019) 2:69 | https://doi.org/10.1038/s42004-019-0168-6 | www.nature.com/commschem 7 ARTICLE COMMUNICATIONS CHEMISTRY | https://doi.org/10.1038/s42004-019-0168-6 used to assemble pharmaceuticals including SC-560, mavacoxib, fused scaffolds scope (>25 examples). Considering the important and celecoxib49,50. role of COXs inhibition in antiplatelet therapy, we also developed drug-like platelet aggregation inhibitor synthesis using TAC 3-trifluormethylpyrazole synthesis in pharmaceutical applica- protocol. Notably, further applications of Lewis base catalysis for tions. To further evaluate the pharmaceutical implication of the synthesis of related heterocycles will be reported in due this TAC protocol, the 3-trifluormethylpyrazole synthesis in course. drug discovery was also explored. Recently, diaryl-3- trifluormethylpyrazoles have been successfully screened as Methods COXs inhibitor and exerted selectively COX-1 or COX-2 General procedure for the TAC reactions. A dried Schlenk tube is charged with the alkynes 1 (0.30 mmol, 1.0 equiv.), CF3CHN2 2 (1.2 mmol, 4.0 equiv., and1.5 M inhibition, such as SC-560 (selective COX-1 inhibitor). Con- in toluene), and 0.4 mL 1,4-dioxane. Subsequently, DBU (0.06 mmol, 20 mol%) is sidering the potential combined pharmacological effects of successively added. The resulting yellow solution is stirred at room temperature COXs inhibition and P2Y12-receptor antagonism in antiplate- until the reaction is complete (as monitored by TLC). After the solvent is evapo- let therapy51,52, we showcased the structural features of clopi- rated under reduced pressure, the crude product is purified via flash chromatography (pentane/ethyl acetate 10:1–3:1) to give the 3-trifluormethylpyrazoles 3 as dogrel (P2Y12-receptor antagonist) and SC-560 (COX-1 white solids. A similar procedure was used for the synthesis of 3- inhibitor) that allow novel analogs to mimic theirs functions trifluormethylpyrazole derivatives 5, 7, 9, and 10. and then 3-trifluormethylpyrazole scaffold was embedded onto thienopyridine ring via TAC strategy (Fig. 6a, b) (see Supple- Synthetic applications. The procedures for 10-g-scale synthesis, pharmaceuticals mentary Methods and Supplementary Fig. 4). Later, we synthesis, and drug-like molecules development are available in the Supplemen- downloaded the crystal structure of COX-1 (3N8Y) from PDB tary Methods and Supplementary Figs. 1–4. andusedSurflex-dock (SYBYL X-2.0) to complete molecular docking53. It was found that the N-atom of the pyrazole ring Platelet aggregation inhibition assay. Please see Supplementary Methods and Supplementary Fig. 5. (10a, 10b) showed H-bonding with Tyr385 (distance N–H = 2.6 Å for 10a and 2.4 Å for 10b, respectively), while 10c H- NMR spectra. 1H, 13C, and 19F NMR spectra of purified compounds are available bonded to Tyr385 and Ser530 (key amino-acid residues in in Supplementary Figs. 8–229. COX-1 binding pocket) via the F-atom of the CF3 group (dis- – = tance F H 2.2 Å for Tyr385 and 2.0 Å for Ser530, respec- Crystallography. X-ray crystallographic CIF files for compounds 5a and 5k are tively). In addition, the N-atom of the pyrazole ring (10b, 10c) available in Supplementary Data 1, 2, and Supplementary Figs. 230, 231. also showed H-bonding with Ser530 (distance N–H = 2.6 Å for 10b and 2.8 Å for 10c, respectively) (Fig. 6c). These results Data availability suggested that the synthetic analogs had potential COX-1 The authors declare that all the data supporting the findings of this study are available inhibition (see Supplementary Table 2). To prove the reason- within the paper and its supplementary information files, or from the corresponding able of the docking model and possible pharmacological author upon request. The X-ray crystallographic coordinate for structure reported in this article has been deposited at the Cambridge Crystallographic Data Center (CCDC properties, the platelet aggregation inhibition assay in vitro was 1886034, CCDC 1907151). These data could be obtained free of charge from the evaluated in parallel with aspirin. We found 10c could dra- Cambridge Crystallographic Data Center via www.ccdc.cam.ac.uk/data_request/cif. matically inhibit platelet aggregation which induced by arachidonic acid (Fig. 6d) (see Supplementary Fig. 5). The result Received: 12 February 2019 Accepted: 21 May 2019 indicated the promise of 10c for future pharmaceutical applications.

Mechanistic proposal. A postulated mechanism for this Lewis base catalysis via the triazene intermediate is proposed References 54,55 (Fig. 7a) . A deuterium-labeling experiment was conducted 1. Janin, Y. L. Preparation and chemistry of 3/5-halogenopyrazoles. Chem. Rev. ′ ′ – using deuterated alkyne (1a ) with CF3CHN2. Compound 3a 112, 3924 3958 (2012). with 90% deuterium incorporation onto the pyrazole ring was 2. Ansari, A., Ali, A., Asif, M. & Shamsuzzaman Review: biologically active – obtained, which supports our proposed 1,3-H shift. To confirm pyrazole derivatives. New J. Chem. 41,16 41 (2017). 3. Lamberth, C. Pyrazole chemistry in crop protection. Heterocycles 71, the DBU activation of CF3CHN2 to form triazene intermediate in 1467–1502 (2007). the cycloaddition reaction, experiments between DBU and 4. Yet, L. Comprehensive heterocyclic chemistry III. 1st edn, Vol. 4, (1–141. CF3CHN2 (in CDCl3) were conducted (Fig. 7c). The postulated Elsevier, Oxford, 2008). intermediate IV was captured from the crude 1H NMR and 5. Zhou, Y. et al. Next generation of fluorine-containing pharmaceuticals, – LC–MS experiments (see Supplementary Figs. 6 and 7), which compounds currently in phase II III clinical trials of major pharmaceutical companies: new structural trends and therapeutic areas. Chem. Rev. 116, further supports the possible mechanism for this Lewis base 422–518 (2016). catalysis. 6. Meyer, F. Trifluoromethyl nitrogen heterocycles: synthetic aspects and potential biological targets. Chem. Commun. 52, 3077–3094 (2016). 7. Mykhailiuk, P. K. Heptafluoroisopropyl diazomethane (i-C3F7CHN2): in situ Discussion generation and synthesis of pyrazoles. Org. Biomol. Chem. 15, 7296–7301 We have developed a TAC reaction to synthesize a series of 3- (2017). trifluormethylpyrazole heterocycles in high to excellent yields. In 8. Penning, T. D. et al. Synthesis and biological evaluation of the 1,5- addition, with the newly explored transformation, the cycload- diarylpyrazole class of cyclooxygenase-2 inhibitors: identification of 4-[5-(4- fl dition strategy is extended to enable LSF of pharmaceuticals and methylphenyl)-3- (tri uoromethyl)-1H-pyrazol-1-yl] benzenesulfonamide (SC-58635, Celecoxib). J. Med. Chem. 40, 1347–1365 (1997). fuse various scaffolds, ranging from clinical drugs and natural 9. Sakya, S. M. et al. 5-Heteroatom substituted pyrazoles as canine COX-2 products to bioactive heterocycles, which may aid the efficiency of inhibitors. part III: molecular modeling studies on binding contribution of 1- lead compound and drug discovery processes. The protocol, (5-methylsulfonyl) pyrid-2-yl and 4-nitrile. Bioorg. Med. Chem. Lett. 17, featured in an operationally simple and environmentally friendly 1067–1072 (2007). manner in comparison to transition-metal catalysis, exhibits a 10. Sun, A., Chandrakumar, N., Yoon, J. J., Plemper, R. K. & Snyder, J. P. Non- nucleoside inhibitors of the measles virus RNA-dependent RNA polymerase broad, structurally diverse substrate scope (>40 examples), and

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complex activity: synthesis and in vitro evaluation. Bioorg. Med. Chem. Lett. 35. Qin, S., Zheng, Y., Zhang, F. G. & Ma, J. A. One-pot cascade transformations 17, 5199–5203 (2007). of zinc trifluorodiazoethylide and α,β-unsaturated enones: access to 11. Harms, J. E., Benveniste, M., Maclean, J. K. F., Partin, K. M. & Jamieson, C. trifluoromethylated polycyclic pyrazolines. Org. Lett. 19, 3406–3409 (2017). Functional analysis of a novel positive allosteric modulator of AMPA 36. Tinoco, A., Steck, V., Tyagi, V. & Fasan, R. Highly diastereo- and receptors derived from a structure-based drug design strategy. enantioselective synthesis of trifluoromethyl-substituted cyclopropanes via Neuropharmacology 64,45–52 (2013). myoglobin-catalyzed transfer of trifluoromethylcarbene. J. Am. Chem. Soc. 12. Norris, T., Colon-Cruz, R. & Ripin, D. H. B. New hydroxy-pyrazoline 139, 5293–5296 (2017). intermediates, subtle regio-selectivity and relative reaction rate variations 37. Chen, Z., Zheng, Y. & Ma, J. A. Use of a traceless activating and directing observed during acid catalyzed and neutral pyrazole cyclization. Org. Biomol. group for the construction of trifluoromethylpyrazoles: one-pot Chem. 3, 1844–1849 (2005). transformation of nitroolefins and trifluorodiazoethane. Angew. Chem. Int. Ed. 13. Singh, S. P., Kumar, V., Aggarwal, R. & Elguero, J. The reaction of aryl and 56, 4569–4574 (2017). heteroarylhydrazines with aryl-trifluoromethyl β-diketones. J. Heterocycl. 38. Guo, R., Lv, N., Zhang, F. G. & Ma, J. A. Zinc-mediated mannich-type – fl β Chem. 43, 1003 1014 (2006). reaction of 2,2,2-tri uorodiazoethane with imines: access to -CF3-amines. 14. Montoya, V. et al. Reaction of 2-hydroxyethylhydrazine with a Org. Lett. 20, 6994–6997 (2018). trifluoromethyl-β-diketone: study and structural characterization of a new 5- 39. Zhang, X. Y. et al. Use of trifluoroacetaldehyde N-tfsylhydrazone as a hydroxy-5-trifluoromethyl-4,5-dihydropyrazole intermediate. J. Fluor. Chem. trifluorodiazoethane surrogate and its synthetic applications. Nat. Commun. 128, 1007–1011 (2007). 10, 284 (2019). 15. Fustero, S. et al. Improved regioselectivity in pyrazole formation through the 40. Wang, Z. F., Herraiz, A. G., del Hoyo, A. M. & Suero, M. G. Generating use of fluorinated alcohols as solvents: synthesis and biological activity of carbyne equivalents with photoredox catalysis. Nature 554,86–91 (2018). fluorinated febufenpyrad analogs. J. Org. Chem. 73, 3523–3529 (2008). 41. Kimball, D. B. & Haley, M. M. Triazenes: a versatile tool in organic synthesis. 16. Mykhailiuk, P. K. et al. Synthesis of trifluoromethyl-substituted proline Angew. Chem. Int. Ed. 41, 3338–3351 (2002). analogues as 19F NMR labels for peptides in the polyproline II conformation. 42. Wang, S., Yang, L. J., Zeng, J. L., Zheng, Y. & Ma, J. A. Silver-catalyzed [3 + 2] Angew. Chem. Int. Ed. 47, 5765–5767 (2008). cycloaddition of isocyanides with diazo compounds: new regioselective access 17. Morandi, B. & Carreira, E. M. Iron-catalyzed cyclopropanation with to 1,4-disubstituted-1,2,3-triazoles. Org. Chem. Front. 2, 1468–1474 (2015). trifluoroethylamine hydrochloride and olefins in aqueous media: in situ 43. Dai, H. X., Stepan, A. F., Plummer, M. S., Zhang, Y. H. & Yu, J. Q. Divergent generation of trifluoromethyl diazomethane. Angew. Chem. Int. Ed. 49, C–H functionalizations directed by sulfonamide pharmacophores: late-stage 938–941 (2010). diversification as a tool for drug discovery. J. Am. Chem. Soc. 133, 7222–7228 18. Morandi, B. & Carreira, E. M. Rhodium-catalyzed cyclopropenation of (2011). alkynes: synthesis of trifluoromethyl-substituted cyclopropenes. Angew. Chem. 44. Wencel-Delord, J. & Glorius, F. C-H bond activation enables the rapid Int. Ed. 49, 4294–4296 (2010). construction and late-stage diversification of functional molecules. Nat. Chem. 19. Morandi, B., Mariampillai, B. & Carreira, E. M. Enantioselective cobalt- 5, 369–375 (2013). catalyzed preparation of trifluoromethyl-substituted cyclopropanes. Angew. 45. Cernak, T., Dykstra, K. D., Tyagarajan, S., Vachalb, P. & Krskab, S. W. The Chem. Int. Ed. 50, 1101–1104 (2011). medicinal chemist’s toolbox for late stage functionalization of drug-like 20. Morandi, B. & Carreira, E. M. Synthesis of trifluoroethyl-substituted ketones molecules. Chem. Soc. Rev. 45, 546–576 (2016). from aldehydes and cyclohexanones. Angew. Chem. Int. Ed. 50, 9085–9088 46. Liu, Z. L. et al. SuFEx click chemistry enabled late-stage drug (2011). functionalization. J. Am. Chem. Soc. 140, 2919–2925 (2018). 21. Liu, C. B. et al. A facile parallel synthesis of trifluoroethyl-substituted alkynes. 47. Pliska, V., Testa, B. & Waterbeemd, H. (eds) Lipophilicity in drug action and Angew. Chem. Int. Ed. 51, 6227–6230 (2012). toxicology (Wiley-VCH, Weinheim, 2008). 22. Chai, Z., Bouillon, J. P. & Cahard, D. Chiral Brønsted acid-catalyzed diastereo- 48. Bhardwaj, A., Kaur, J., Wuest, M. & Wuest, F. In situ click chemistry and enantioselective synthesis of CF3-substituted aziridines. Chem. Commun. generation of cyclooxygenase-2 inhibitors. Nat. Commun. 8, 1 (2017). 48, 9471–9473 (2012). 49. Penning, T. D. et al. Synthesis and biological evaluation of the 1,5- 23. Molander, G. A. & Cavalcanti, L. N. Synthesis of trifluoromethylated diarylpyrazole class of cyclooxygenase-2 inhibitors: identification of 4-[5-(4- ssoxazolidines: 1,3-dipolar cycloaddition of nitrosoarenes, (trifluoromethyl) methylphenyl)-3- (trifluoromethyl)-1H-pyrazol-1-yl] benzenesulfon-amide diazomethane, and alkenes. Org. Lett. 15, 3166–3169 (2013). (SC-58635, Celecoxib). J. Med. Chem. 40, 1347–1365 (1997). 24. Argintaru, O. A., Ryu, D., Aron, I. & Molander, G. A. Synthesis and 50. Wang, Y. D., Han, J., Chen, J. & Cao, W. G. An efficient route to 3- applications of α-trifluoromethylated alkylboron compounds. Angew. Chem. trifluoromethylpyrazole via cyclization/1,5-H shift and its applications in the Int. Ed. 52, 13656–13660 (2013). synthesis of bioactive compounds. Tetrahedron 71, 8256–8262 (2015). 25. Li, F., Nie, J., Sun, L., Zheng, Y. & Ma, J. A. Silver-mediated cycloaddition of 51. Warner, T. D., Nylander, S. & Whatling, C. Anti-platelet therapy: cyclo- alkynes with CF3CHN2: highly regioselective synthesis of 3- oxygenase inhibition and the use of aspirin with particular regard to dual anti- trifluoromethylpyrazoles. Angew. Chem. Int. Ed. 52, 6255–6258 (2013). platelet therapy. Br. J. Clin. Pharmacol. 72, 619–633 (2011). ’ 26. Li, T. R. et al. Synthesis of CF3-containing 3,3 -cyclopropyl spirooxindoles by 52. Zerna, C., Thomalla, G., Campbell, B. C. V., Rha, J. H. & Hill, M. D. Current sequential [3 + 2] cycloaddition/ring contraction of ylideneoxindoles with practice and future directions in the diagnosis and acute treatment of 2,2,2-trifluorodiazoethane. J. Org. Chem. 79, 2296–2302 (2014). ischaemic stroke. Lancet 392, 1247–1256 (2018). 27. Xiong, H. Y. et al. Copper-catalyzed one-pot denitrogenative- 53. Sidhu, R. S., Lee, J. Y., Yuan, C. & Smith, W. L. Comparison of dehydrogenative-decarboxylative coupling of β-ketoacids with cyclooxygenase-1 crystal structures: cross-talk between monomers comprising trifluorodiazoethane: facile access to trifluoromethylated aldol products. cyclooxygenase-1 homodimers. Biochemistry 49, 7069–70792 (2010). Chem. Eur. J. 20, 8325–8332 (2014). 54. Aricò, F., Evaristoa, S. & Tundoa, P. Synthesis of five- and six-membered 28. Mao, H. et al. Organocatalytic one-pot synthesis of highly substituted heterocycles by dimethyl carbonate with catalytic amounts of nitrogen bicyclic pyridazines from Morita-Baylis-Hillman carbonates and diazo compounds. bases. Green Chem. 17, 1176–1185 (2015). Chem. Eur. J. 20, 2454–2458 (2014). 55. Krishna, P. R. & Prapurna, Y. L. DABCO catalyzed facile synthesis of highly 29. Molander, G. A. & Ryu, D. Diastereoselective synthesis of vicinally bis functionalized pyrazolines from Baylis–Hillman acetates and ethyl (trifluoromethylated) alkylboron compounds through successive insertions of diazoacetate. Tetrahedron Lett. 51, 6507–6510 (2010). 2,2,2-trifluorodiazoethane. Angew. Chem. Int. Ed. 53, 14181–14185 (2014). 30. Zhang, F. G., Wei, Y., Yi, Y. P., Nie, J. & Ma, J. A. Regioselective cycloaddition fl fi of tri uorodiazoethane with electron-de cient allenic esters and ketones: Acknowledgements access to CF -substituted pyrazolines and pyrazoles. Org. Lett. 16, 3122–3125 3 We thank Prof. Dr Dieter Enders for the valuable suggestions. We are also grateful for (2014). financial support from the National Natural Science Foundation of China (81602977), 31. Guo,R.,Zheng,Y.&Ma,J.A.Electrophilicreactionof2,2,2-trifluorodiazoethane National Science and Technology Major Project (no. 2018ZX09711001-009-001), the with the in situ generated N-heterocyclic carbenes: access to N-aminoguanidines. CAMS Innovation Fund for Medical Sciences (2016-I2M-1-012; 2017-I2M-1-013; 2018- Org. Lett. 18, 4170–4173 (2016). I2M-HL-010), the Science and Technology Development Project of Jilin Province of 32. Luo, H., Wu, G., Zhang, Y. & Wang, J. Silver(I)-catalyzed N- China (20190304050YY), and YESS (2017QNRC001). trifluoroethylation of anilines and O-trifluoroethylation of amides with 2,2,2- trifluorodiazoethane. Angew. Chem. Int. Ed. 54, 14503–14507 (2015). 33. Hyde, S. et al. Copper-catalyzed insertion into heteroatom-hydrogen bonds with trifluorodiazoalkanes. Angew. Chem. Int. Ed. 55, 3785–3789 (2016). Author contributions 34. Arkhipov, A. V., Arkhipov, V. V., Cossy, J., Kovtunenko, V. O. & Mykhailiuk, S.P.L. and H.Z. performed the experiments and analyzed the experimental data with P. K. Unexpected reactivity of trifluoromethyl diazomethane (CF3CHN2): contributions of X.Y., Y.X., H.J.Z., M.W., and Z.R.D run the molecule docking, H.T.L. electrophilicity of the terminal N-atom. Org. Lett. 18, 3406–3409 (2016). and G.B.S. developed the in vitro experiments, X.J.G. carried out the X-ray structure

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determinations. L.W. directed the investigations, X.B.S. and L.W. prepared the manu- Open Access This article is licensed under a Creative Commons script with contributions of S.P.L. and H.Z. Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give Additional information appropriate credit to the original author(s) and the source, provide a link to the Creative Supplementary information accompanies this paper at https://doi.org/10.1038/s42004- Commons license, and indicate if changes were made. The images or other third party ’ 019-0168-6. material in this article are included in the article s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the Competing interests: The authors declare no competing interests. article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from Reprints and permission information is available online at http://npg.nature.com/ the copyright holder. To view a copy of this license, visit http://creativecommons.org/ reprintsandpermissions/ licenses/by/4.0/.

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