Corrections

BIOCHEMISTRYCorrection for “Confinement of caspase-12 proteolytic activity toautoprocessing,” by Sophie Roy, Jeffrey R. Sharom, CarolineHoude, Thomas P. Loisel, John P. Vaillancourt, Wei Shao, MayaSaleh, and Donald W. Nicholson, which appeared in issue 11,March 18, 2008, of Proc Natl Acad Sci USA (105:4133–4138; firstpublished March 10, 2008; 10.1073/pnas.0706658105).The authors note that Fig. 6 appeared incorrectly. There was

an error in the alignment of the molecular mass markers, andminor adjustments have been made to the assignment of caspase-12 bands. The authors also note that the source of the anti-caspase-12 antibody for Fig. 6 was Sigma (clone 14F7). Thiserror does not affect the conclusions of the article. The correctedfigure and its corresponding legend appear below.

www.pnas.org/cgi/doi/10.1073/pnas.1302753110

CHEMISTRYCorrection for “Direct asymmetric vinylogous Michael additionof cyclic enones to nitroalkenes via dienamine catalysis,” byGiorgio Bencivenni, Patrizia Galzerano, Andrea Mazzanti,Giuseppe Bartoli, and Paolo Melchiorre, which appeared in issue48, November 30, 2010, of Proc Natl Acad Sci USA (107:20642–20647; first published June 21, 2010; 10.1073/pnas.1001150107).The authors note that they incorrectly assigned the structure

of the reaction product reported in Scheme 4. The publishedstructure represents the γ-aminated adduct 8, when it shouldinstead be the α-analogue arising from an α-site selective path-way. As a result of this, Scheme 4 and its related commentsshould be removed from the article.On page 20642, left column, within the Abstract, lines 16–18,

“Finally, we describe the extension of the dienamine catalysis-induced vinylogous nucleophilicity to the asymmetric γ-aminationof cyclohexene carbaldehyde” should be removed from the article.On page 20645, right column, third full paragraph, lines 1–8,

to page 20646, left column, first paragraph, lines 1–2, “Finally, toexplore the potential of the chiral primary amine-induced vi-nylogous nucleophilicity, we wondered whether this unique re-activity concept may be translated to an aldehyde derivativeadorned with a six-membered ring scaffold, reminiscent of theβ-substituted cyclohexanone framework. Although the vinylogousMichael addition of 1-cyclohexene-1-carboxaldehyde 7 to ni-trostyrene 2a did not proceed at all, the combination with tert-butylazodicarboxylate under the catalysis of A furnished theγ-amination product 8 with perfect regio- and enantioselectivity(Scheme 4)” should be removed from the article.These errors do not affect the conclusions of the article of the

vinylogous Michael addition of cyclic enones to nitroalkenes. Theability of primary amine catalysis to address the synthetic issueconnected with the enantioselective carbon–carbon bond forma-tion gamma to a carbonyl group, promoting vinylogous nucleo-philicity upon selective activation of unmodified cyclic unsaturatedketones, is fully supported by the separated results presented inTables 1, 2, and 3, and Schemes 2 and 3.

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Fig. 6. Procaspase-12 forms a complex with caspase-1 and is partially au-toprocessed in the complex. HEK293T cells were cotransfected with expres-sion vectors harboring Flag-tagged procaspase-1 (all lanes) plus eitherprocaspase-12 (lanes 1 and 4) or the catalytically incapacitated C299A mutant(lanes 2 and 5). After 24 h, cells were harvested and lysed. One tenth of thelysate was directly applied to SDS/PAGE (lanes 1 and 2), and the remainderwas immunoharvested with antibodies directed against the caspase-1 Flagepitope tag (lanes 4 and 5; lane 3 was processed in the same way, except thatonly lysis buffer was used). Immunoblotting for the large subunit (p20) ofcaspase-12 revealed that procaspase-12 and the resulting autocleavageproduct were both immunoharvested with caspase-1. The asterisk indicatesa band of unknown identity that is detected by prebleed control serum.

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MICROBIOLOGYCorrection for “Commensal bacteria play a role in matingpreference of Drosophila melanogaster,” by Gil Sharon, DanielSegal, John M. Ringo, Abraham Hefetz, Ilana Zilber-Rosenberg,and Eugene Rosenberg, which appeared in issue 46, November16, 2010, of Proc Natl Acad Sci USA (107:20051–20056; first pub-lished November 1, 2010; 10.1073/pnas.1009906107).The authors note the following: “The mating frequencies

reported in Table 1 of this paper did not follow a multinomialdistribution, making the statistical analysis inapplicable. Thisproblem was obviated by considering only the first matingsin each experimental unit and computing odds ratios. Aftersubmitting the paper, we continued to perform experiments

identical in design to those we reported. In the table below,we combined the results of those additional replicate experi-ments with those already reported. From the new analysis, wenow find that experiment 4, in which flies were infected witha mixture of Lactobacillus spp., assortative mating was notrestored. Otherwise, the conclusions of the article were notchanged by our reanalysis. We acknowledge the statistical ad-vice of Dan Yekutieli and thank Tal Lahav for calculating theodds ratios and their 95% confidence intervals, and for per-forming the chi-squared tests presented in the correctedTable 1.”The corrected Table 1 appears below.

34. Mantel N, Haenszel W (1959) Statistical aspects of the analysis of data from retrospective studies of disease. J Natl Cancer Inst 22(4):719–748.35. Cochran WD (1954) Some methods for strengthening the common χ2 tests. Biometrics 10:417–451.

www.pnas.org/cgi/doi/10.1073/pnas.1302326110

MEDICAL SCIENCESCorrection for “Interaction of intracellular β amyloid peptidewith chaperone proteins,” by Virginia Fonte, Vadim Kapulkin,Andrew Taft, Amy Fluet, David Friedman, and ChristopherD. Link, which appeared in issue 14, July 9, 2002, of Proc NatlAcad Sci USA (99:9439–9444; first published June 27,2002; 10.1073/pnas.152313999).The authors note that the author name Vadim Kapulkin should

instead appear as Wadim Jan Kapulkin. The corrected author lineappears below. The online version has been corrected.

Virginia Fonte, Wadim Jan Kapulkin, Andrew Taft,Amy Fluet, David Friedman, and Christopher D. Link

www.pnas.org/cgi/doi/10.1073/pnas.1302545110

SYSTEMS BIOLOGYCorrection for “Expanded methyl-sensitive cut counting revealshypomethylation as an epigenetic state that highlights functionalsequences of the genome,” by Alejandro Colaneri, Nickolas Staffa,David C. Fargo, Yuan Gao, Tianyuan Wang, Shyamal D. Peddada,and Lutz Birnbaumer, which appeared in issue 23, June 7, 2011,of Proc Natl Acad Sci USA (108:9715–9720; first publishedMay 20, 2011; 10.1073/pnas.1105713108).The authors note that, within the corresponding author foot-

note on page 9715, the email address “colaneria@niehs.nih.gov”should instead appear as “acolaneri_2000@yahoo.com.ar”.

www.pnas.org/cgi/doi/10.1073/pnas.1302473110

Table 1. The role of bacteria in diet-induced mating preference of D. melanogaster

Experiment Fly treatment* N† OR‡ 95% CI P value‡

1 Starch-grown × CMY-grown 18 3.21 2.14–4.81 1.8 × 10−8

2 Experiment 1 after antibiotics 11 1.04 0.63–1.71 0.98883 Experiment 2 after infection of starch-grown flies with homologous bacteria§ 6 2.68 1.40–5.11 0.04774 Experiment 3 with Lactobacillus spp. replacing homologous bacteria 4 1.76 0.74–4.19 0.29125 Experiment 3 with Lactobacillus plantarum replacing homologous bacteria 7 2.14 1.35–3.39 0.00196 Infection control (no added bacteria) 4 1.26 0.53–3.00 0.7712

*After all treatments, the flies were grown for one generation in CMY medium before performing the mating preference test.†N is the number of replicate experiments.‡Cochran-Mantel-Haenszel Odds Ratio and P value are from the Cochran-Mantel-Haenszel Chi-squared test (34, 35).§Antibiotic-treated starch- and CMY-grown flies were infected with bacteria isolated from their respective growth medium (before antibiotic treatment).

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Direct asymmetric vinylogous Michael addition ofcyclic enones to nitroalkenes via dienamine catalysisGiorgio Bencivennia, Patrizia Galzeranoa, Andrea Mazzantia, Giuseppe Bartolia, and Paolo Melchiorreb,c,1

aDipartimento di Chimica Organica “A. Mangini,” Università di Bologna, Viale Risorgimento 4, 40136 Bologna, Italy; bInstitució Catalana de Recerca iEstudis Avançats, Passeig Lluís Companys 23-08010 Barcelona, Spain; and cInstitute of Chemical Research of Catalonia, Avenida Països Catalans 16-43007Tarragona, Spain

Edited by David W. C. MacMillan, Princeton University, Princeton, NJ, and accepted by the Editorial Board May 29, 2010 (received for review February 1, 2010)

In spite of the many catalytic methodologies available for theasymmetric functionalization of carbonyl compounds at their αand β positions, little progress has been achieved in the enantiose-lective carbon–carbon bond formation γ to a carbonyl group. Here,we show that primary amine catalysis provides an efficient way toaddress this synthetic issue, promoting vinylogous nucleophilicityupon selective activation of unmodified cyclic α,β-unsaturatedketones. Specifically, we document the development of the unpre-cedented direct and vinylogous Michael addition of β-substitutedcyclohexenone derivatives to nitroalkenes proceeding under die-namine catalysis. Besides enforcing high levels of diastereo- andenantioselectivity, chiral primary amine catalysts derived fromnatural cinchona alkaloids ensure complete γ-site selectivity: Theresulting, highly functionalized vinylogous Michael adducts,having two stereocenters at the γ and δ positions, are synthesizedwith very high fidelity. Finally, we describe the extension of thedienamine catalysis-induced vinylogous nucleophilicity to theasymmetric γ-amination of cyclohexene carbaldehyde.

asymmetric synthesis ∣ Michael reaction ∣ organocatalysis

The stereoselective functionalization of carbonyl compounds,particularly while concomitantly forming a new carbon–

carbon bond, represents one of the more efficient and potentchemical ways to produce valuable chiral molecules. This targethas inspired generations of synthetic chemists to design uniquechiral catalysts able to enantioselectively forge a stereocenter αor β to a carbonyl group (1–3). In this context, intense investiga-tions into the aldol (4, 5) and Michael reactions (6, 7) have madethem invaluable tools in modern organic chemistry. Recently, theclassical organometallic-based approach has been enriched by thepossibility of using chiral primary or secondary amines as efficientcatalysts for the asymmetric functionalization of carbonyl com-pounds. This strategy is known as “asymmetric aminocatalysis”(8–11). Among the many advantages of this synthetic approach(12), one of its more attractive features is the ability to directlygenerate, in situ, the catalytically active intermediates fromunmodified carbonyl compounds (13, 14).

Both metal- and organic-based approaches have achievedlevels of reliability such that synthetic chemists can now addresseven the most daunting issues connected with the asymmetric cat-alytic functionalization of carbonyl compounds at their α and βpositions. In contrast, little progress has been reported in the cor-responding enantioselective carbon–carbon bond formation γ toa carbonyl group (15). Among the few useful approaches devisedto date, the concept of vinylogous nucleophilicity is the mostpowerful (16–19). Formulated by Fuson in 1935 as the transmis-sion of electronic effects through a conjugated π system (20), thisprinciple accounts for the use of γ-enolizable α,β-unsaturated car-bonyl compounds as precursors of nucleophilic dienolate equiva-lents, formally inverting the usual reactivity of this compoundclass. Vinylogous processes offer an efficient entry onto functio-nalized building blocks having high level of structural complexity;however, designing asymmetric catalytic versions is not simple.Indeed, every approach to vinylogous reactions overlays the

challenge of site-selectivity onto the already present issue ofstereo-selectivity. In general, the critical regiochemical issuecan be addressed by judiciously preparing preformed, stabledienolate equivalents. This strategy has been successfully appliedto asymmetric vinylogous aldol (17, 21), Mannich (18, 22), andMichael reactions (19, 23). Avoiding the stoichiometric preacti-vation of the vinylogous nucleophilic components would logicallyimprove this approach, particularly from the standpoint of atomeconomy (24). However, examples of direct, catalytic, and asym-metric vinylogous reactions are rare: Recently, Trost and Hitcereported on the direct vinylogous Michael addition of 2(5H)-furanone to nitroalkenes under dinuclear zinc catalysis (25),whereas chiral Brønsted base catalysis proved successful to acti-vate specific reactive alkenes, such as α,α-dicyano olefins, towardvinylogous nucleophilicity (26).

Within this context, we wondered whether asymmetric amino-catalysis could solve the challenge of a direct vinylogous nucleo-philic addition of unmodified γ-enolizable carbonyl compounds.We were inspired by a recent report by Jørgensen and coworkerson the direct, enantioselective γ amination of α,β-unsaturatedaldehydes using azodicarboxylates as the electrophilic nitrogensource (27). The process is based on a unique activation mode,dienamine catalysis, which exploits the condensation of a chiralsecondary amine catalyst with β-aliphatic-substituted unsaturatedcompounds. This condensation leads to the formation of theexpected electrophilic iminum ion intermediate, which is in equi-librium with an electron-rich dienamine intermediate. The diena-mine, exploiting its γ-nucleophilic character, actually representsthe catalytic active species. In principle, the ability to promote thein situ formation of a dienamine from γ-enolizable carbonyl com-pounds while enforcing γ-site selectivity during the nucleophilicpath might offer a unique and potent way to design direct viny-logous processes (Fig. 1).

Fig. 1. Dienamine catalysis and the concept of vinylogy; Elec, electrophile.

Author contributions: P.M. designed research; G. Bencivenni and P.G. performed research;G. Bartoli contributed new reagents/analytic tools; A.M. analyzed data; and P.M. wrotethe paper.

The authors declare no conflict of interest.

This article is a PNAS Direct Submission. D.W.C.M. is a guest editor invited by theEditorial Board.

Data deposition: The atomic coordinates have been deposited in the Cambridge StructuralDatabase, Cambridge Crystallographic Data Centre, Cambridge CB2 1EZ, United Kingdom(CSD reference nos. 763174, 763175, and 763176).1To whom correspondence should be addressed. E-mail: pmelchiorre@iciq.es.

This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10.1073/pnas.1001150107/-/DCSupplemental.

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In spite of the potential to address important synthetic issues,such as the effective catalytic generation of a new carbon–carbonbond and a new stereocenter γ to a carbonyl group, dienaminecatalysis has found limited application (28, 29). Accordingly, arecently published perspective on the advent of organocatalysisdid not include dienamine catalysis among the generic modesof activation and induction (30). This lack was probably due tothe fact that γ-amination of unsaturated aldehydes was originallyproposed to follow a [4þ 2] cycloaddition path (27), instead of amore generalizable nucleophilic addition manifold. Moreover,some recent studies suggest that chiral secondary amines, suchas proline (31) and its derivatives (32, 33), can activate γ-enoliz-able unsaturated aldehydes toward the formation of the diena-mine intermediate, but generally promoting an α-site-selectivealkylation step via enamine catalysis in the presence of suitableelectrophiles. Nonetheless, a recent inspiring report by Christ-mann and coworkers has highlighted the potential of dienaminecatalysis for enforcing a direct nucleophilic γ-addition path, albeitin an intramolecular way (34).

Here, we show that dienamine catalysis, induced by chiral pri-mary amines, can efficiently promote the direct, intermolecularvinylogous Michael addition of unmodified β-substituted cyclo-hexenone derivatives to nitroalkenes, imparting high levels ofdiastereo- and enantioselectivity, and ensuring exclusive γ-site se-lectivity. Notably, the two stereocenters at the γ and δ positions ofthe carbonyl moiety are formed with very high fidelity (Scheme 1).

Results and DiscussionBackground. Our laboratory and others, independently, have re-cently introduced 9-amino(9-deoxy)epicinchona alkaloids(Fig. 2), chiral primary amines easily derived from naturalsources, as general and effective catalysts for a wide variety ofasymmetric α and β functionalizations of ketones (35, 36). Wehave further demonstrated the ability of catalyst A, derived fromhydroquinine, to combine orthogonal aminocatalytic modes(iminium and enamine activations) into onemechanism, thus pro-moting cascade reactions with α,β-unsaturated ketones (37, 38)and even with the challenging α,β-disubstituted enals (39).

Next, we decided to investigate the behavior of this versatilecatalyst in the context of the elusive γ-site activation of unmodi-fied enones, in order to design a dienamine-catalyzed direct,vinylogous Michael reaction. From the outset, we were fullyaware of the inherent difficulties of our target, because thepresence of multiple potential sites of enolization has greatlyhampered the use of α,β-unsaturated ketones in even the stoi-chiometric version of vinylogous reactions (40). We decided toattack this problem by selecting β-substituted cyclohexenone de-rivatives as a model substrate. At first glance, this compound classseems highly challenging, because it further enhances the task ofregioselectivity (Fig. 3). The condensation of a chiral primaryamine catalyst with cyclic enones would lead to the formationof the iminium ion, which could equilibrate, upon selective α′-de-protonation or γ- and γ′-deprotonation, with the kinetic cross-conjugated dienamine intermediate I or the thermodynamicextended dienamines II and III, respectively. Although the for-mer would open an avenue for α′-site alkylation (path a in Fig. 3),the extended dienamines could be alkylated at three differentpositions, namely α, γ, and even γ′, behaving as d2 or d4 reagents(paths b, d, and c, respectively).

Despite the increased complexity posed by β-substituted cyclicenones, our choice was motivated by a variety of considerations.Our experience in designing of cascade reactions (37, 38) stronglyindicated that, when reacted with acyclic, linear α,β-unsaturatedketones, catalyst A easily facilitates the equilibrium between theiminium ion and the nucleophilic cross-conjugated dienamine in-termediate of type I, selectively directing the reaction manifoldtoward an α′-site alkylation (path a in Fig. 3). We thus consideredusing cyclohexenone derivatives to coax the regiocontrolled for-mation of extended dienamines within a more thermodynamicallyfavored six-membered ring scaffold. This idea is based upon re-lated enolization studies demonstrating that, under certain con-ditions, the selective formation of the thermodynamic exo-cyclicenolate of type III is strongly favored over both the endo-isomerand the kinetic cross-conjugated dienolate (41–45). This inherentbehavior of β-substituted cyclohexenones has found a wonderfulapplication in the conceptually unique, direct, vinylogous aldolreaction reported by Yamamoto and coworkers (46, 47): Thecomplete γ-site selectivity induced by the nonchiral, bulky alumi-num-based catalyst has been rationalized on the basis of catalyststeric effects (48, 49), which inhibit base deprotonation at the α′position, whereas thermodynamic factors account for the forma-tion of the reactive exo-dienolate isomer of the β-methyl 2-cyclo-hexen-1-one (46). On these grounds, we considered the unique

Scheme 1. The direct vinylogous Michael addition developed in this study.

Fig. 2. Primary amine catalysts used within this study.

Fig. 3. Challenges arising from the site-selective formation of dienamines.

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ability of catalyst A, in combination with an acidic cocatalyst, toperturb the iminium-dienamine equilibrium, taken together withthermodynamic factors, that may govern the regioselective for-mation of the exo-cyclic dienamine intermediate. We wonderedif this ability might be exploited to develop the challengingγ-site-selective, direct stereoselective addition of β-methyl2-cyclohexen-1-one to nitrostyrene derivatives (following the nu-cleophilic path d in Fig. 3). Theoretical calculations accountingfor a thermodynamically driven site-selective formation of adienamine of type III are reported in the SI Appendix.

Organocatalytic Vinylogous Michael Addition. The vinylogousMichael addition under dienamine catalysis was first evaluatedby mixing 2 equivalents of β-methyl 2-cyclohexen-1-one 1 and ni-trostyrene 2a in toluene (1 M, 48 h, 40 °C). In accordance withourmechanistic postulate, the chiral primary amineA (20 mol%),in combination with 30 mol % of 2-fluorobenzoic acid as the co-catalyst, directed the reaction manifold toward a γ-site-selectiveaddition, leading to compound 3a as the unique product of theprocess and with a good level of enantiocontrol (Table 1, entry 1).Examination of the reaction media revealed that the catalyticprocess was greatly influenced by polarity, with solvents with ahigh dielectric constant strongly affecting both reactivity and en-antioselectivity (Table 1, entries 1–5). The nature of the acidiccocatalysts was also a crucial parameter, with stronger acids lead-ing to worse results (Table 1, entries 1, 6, and 7). This evidencesuggests that fine tuning the carboxylic acid cocatalyst is essentialto modulate the perturbation of the delicate equilibrium betweeniminium ion, cross-conjugated and extended dienamine inter-mediates during the reaction.

These results further consolidate A as a general, highly versa-tile catalyst for the activation of keto compounds, even towardvinylogous nucleophilicity. However, we were not satisfied withthe level of stereocontrol in the present chemistry. We speculatedthat using a bifunctional catalyst capable of simultaneouslyactivating both the electrophilic and nucleophilic componentsmight lead to higher catalytic activity and, more importantly,to better stereocontrol (50). To this end, we tested the potentialof 6′-hydroxy-9-amino-9-deoxyepiquinine B to synergistically and

productively bind the two reaction partners of the vinylogous Mi-chael addition. B has recently been introduced by Chen et al. asan efficient bifunctional catalyst of the asymmetric 1,3-dipolarcycloaddition of cyclic enones (51). Catalyst B greatly improvedthe enantioselectivity as well as the reaction rate of the vinylogousaddition of 1 to 2a while maintaining a complete γ-selectivity(Table 1, entry 8). The enhanced catalytic activity allowed usto lower the catalyst loading to 10 mol % (Table 1, entry 9), de-lineating a more practical synthetic protocol (extensive optimiza-tion studies can be found within the SI Appendix).

The best result was obtained with the catalytic salt made bycombining B (10 mol %) and 2-fluorobenzoic acid (20 mol %)in toluene (0.2 M). These conditions were selected to examinethe scope of the vinylogous Michael addition by evaluating avariety of nitroalkenes (Table 2). Different substituents at the aro-matic moiety of β-nitrostyrene derivatives were well-tolerated, re-gardless of their electronic properties, because the correspondingadducts 3 were obtained in good to high yield and almost perfectstereocontrol (enantiomeric excesses ranging from 95% to 98%).The pseudoenantiomeric catalyst C, derived from quinidine,accounted for the possibility of accessing both antipodes of theproducts (Table 2, entries 2 and 10).

As limitations of the direct vinylogous Michael reactions, ali-phatic nitroalkenes did not react under the described conditions.Moreover, modifying the cyclic scaffold of the nucleophilic com-ponent (i.e., 3-methyl 2-cyclopenten-1-one) resulted in a com-plete loss of reactivity, a result that highlights how strongly thecyclic scaffold geometry influences (and drives) the selective for-mation of the thermodynamic, extended dienamine intermediate.

The absolute configuration of the stereogenic center was un-ambiguously determined to be R by anomalous dispersion X-raycrystallographic analysis of the bromide derivative 3d. The abilityof the catalyst to communicate its inherent stereochemical infor-mation while forging the new stereocenter at the δ position,several atoms apart from the catalyst binding point within thecovalent dienamine intermediate, seemed noteworthy to us. Itis also intriguing to consider how primary amine catalysis canimpart unique mechanistic pathways, thus complementing and

Table 1. Optimization studies of the vinylogous Michael additionunder dienamine catalysis

Entry Amine Acid Solvent Conversion, %* ee, %†

1 A 2F-C6H4CO2H Toluene >95 822 A 2F-C6H4CO2H CHCl3 52 793 A 2F-C6H4CO2H THF 30 824 A 2F-C6H4CO2H MeOH <5 —5 A 2F-C6H4CO2H MeCN 12 —6 A TFA Toluene <5 —7 A 2NO2-C6H4CO2H Toluene 45 808 B 2F-C6H4CO2H Toluene >95 989 B 2F-C6H4CO2H Toluene >95 (77)‡ 98§

TFA, trifluoroacetic acid; reactions carried out using two equivalents ofenone 1*Conversion determined by 1H NMR analysis of the crude mixture; onlyproduct 3a, derived from a γ-site-selective alkylation step, has alwaysbeen detected.

†The enantiomeric excess (ee) was determined by HPLC analysis on chiralstationary phases.

‡Number in parentheses refers to yield of the isolated compound 3a.§Reaction performed in the presence of 10 mol % of B and 20 mol % of2F-C6H4CO2H and using ½2�0 ¼ 0.2 M

Table 2. Vinylogous Michael addition: Scope of the nitrostyrenederivatives

Entry Catalyst R 3 Yield, %* ee, %†

1 B Ph a 77 982 C Ph a 55 95‡

3 B 4-MeO-Ph b 70 974 B 4-Me-Ph c 70 985 B 4-Br-Ph d 73 976 B 2-Cl-Ph e 84 967 B 2-F-Ph f 83 978 B 2-thiophenyl g 68 989 B 2-OBn-Ph h 87 9710 C 2-OBn-Ph h 68 96‡

Bn, benzyl; reactions carried out using two equivalents of enone 1. 1H NMRanalysis of the crude mixture indicated a highly γ-site-selective alkylationpathway, other products arising from different reaction manifolds (e.g.,α′-alkylation under enamine catalysis, see ref. 52) being sporadicallydetected in negligible amounts.*Yield of the isolated compounds 3.†The enantiomeric excess (ee) was determined by HPLC analysis on chiralstationary phases.

‡The opposite (S) enantiomer has been obtained using catalyst C.

20644 ∣ www.pnas.org/cgi/doi/10.1073/pnas.1001150107 Bencivenni et al.

enriching the established reactivity profile of secondary aminecatalysis. Within this context, a recent report has demonstratedthat a chiral secondary amine behaves in a completely differentway when exposed to the very same reagents combination, namelyβ-methyl cyclohexenone and nitrostyrene, catalyzing a Diels-Alder reaction between 1 and 2a via the selective formation ofcross-conjugated dienamine of type I (52).

Our vinylogous protocol could be successfully extended to aβ,β-disubstituted nitrostyrene, leading to the stereocontrolledgeneration of compound 4 having an all-carbon quaternary(53) stereocenter (Scheme 2). Moreover, a different class ofMichael acceptor has proven to be a viable component of thedienamine-catalyzed direct vinylogous addition. Mixing β-methyl2-cyclohexen-1-one with trans-α-cyanocinnamate under the cata-lysis of A furnished the corresponding vinylogous adduct with twostereogenic centers. As reported in Scheme 3, to avoid theepimerization event, we designed a one-pot vinylogous Michaeladdition/amination tandem sequence, directly leading to com-

pound 5 with complete control over the relative stereochemistryand high enantioselectivity (54).

Next, we studied the possibility of forging two contiguousstereogenic centers at the γ and δ positions, examining thevinylogous Michael addition of differently β-substituted cyclohex-anones to a variety of nitrostyrene derivatives under the catalysisof the salt made by combining 20 mol % of the chiral primaryamine B with 30 mol % of 2-fluorobenzoic acid. In line with pre-vious observations, the nature of the carboxylic acid cocatalyststrongly influenced both the reactivity and the stereochemicaloutcome of the process: Carrying out the reaction in the presenceof 40 mol % of salicylic acid afforded higher enantiocontrol andan increased reaction rate, albeit with slightly lower diastereos-electivity (Table 3, entries 1 and 2). Thus, both catalyst salt com-binations, where B is mixed with 2-fluoro- or 2-hydroxybenzoicacids, have been evaluated. Selected results are reported inTable 3. In general, a variety of substrate combinations can berealized, leading to products 6, readily amenable to further func-tionalizations, with high levels of γ-site, diastereo- and enantios-electivity.

Crystals from bromide 6l and from compound 6i were suitablefor X-ray analysis, which established the absolute configuration ofthe vinylogous reaction as well as its anti-stereochemical out-come. Interestingly, the observed sense of relative stereoinduc-tion is not common in the corresponding enamine-catalyzedMichael addition of carbonyls to nitroalkenes, which generallyleads to a syn-relationship (55, 56).

Finally, to explore the potential of the chiral primary amine-induced vinylogous nucleophilicity, we wondered whether thisunique reactivity concept may be translated to an aldehydederivative adorned with a six-membered ring scaffold, reminis-cent of the β-substituted cyclohexanone framework. Althoughthe vinylogous Michael addition of 1-cyclohexene-1-carboxalde-hyde 7 to nitrostyrene 2a did not proceed at all, the combinationwith tert-butylazodicarboxylate under the catalysis of A furnished

Table 3. Stereoselective creation of vicinal stereocenters under dienamine catalysis

Entry Catalyst salt combination R1 R2 6 Yield, %* dr ee, %†

1 a Me Ph a 72 9∶1 922 b Me Ph a 80 6∶1 94§

3 a Me 4-MeO-Ph b 78 10∶1 944 a Me 4-Me-Ph c 76 10∶1 925 b Me 4-Br-Ph d 81 5∶1 936 b Me 4-NO2-Ph e 65 3∶1 957 b Me 2-F-Ph f 80 6∶1 908 b Me 2-thiophenyl g 90‡ 3.3∶1 949 b Bn Ph h 65‡ 7∶1 8510 b allyl Ph i 44 11.5∶1 94§

11 b propyl Ph j 86‡ 11.5∶1 9512 b propyl 4-MeO-Ph k 72 13.5∶1 91§

13 b propyl 4-Br-Ph l 50 10∶1 94§

14 b Ph Ph m 86‡ 2∶1 90§

Bn, benzyl; reactions carried out using two equivalents of enones 1. 1H NMR analysis of the crude mixture indicated a highly γ-site-selective alkylation pathway, other products arising from different reaction manifolds (e.g., α′-alkylation under enamine catalysis, seeref. 52) being sporadically detected in negligible amounts.*Refers to the isolated single, major diastereoisomer.†The enantiomeric excess (ee) was determined by HPLC analysis on chiral stationary phases.‡Refers to the isolated mixture of diastereoisomers.§Enantiomerically pure products obtained after a single crystallization.

Schemes 2 and 3.

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the γ-amination product 8 with perfect regio- and enantioselec-tivity (Scheme 4).

Concluding RemarksThe direct, γ-site-selective addition of β-substituted cyclohexe-none derivatives to nitroalkenes represents one of the few exam-ples of catalytic, asymmetric vinylogous Michael reactions ofunmodified carbonyl compounds. This unprecedented chemicaltransformation affords highly functionalized compounds, havingtwo stereocenters at the γ and δ positions, with high enantiomericpurity. In addition to its synthetic interest, this study confirms theability of chiral primary amine catalysis to impart unique reactiv-ity profiles, thus expanding the potential of asymmetric aminoca-talysis. Specifically, dienamine catalysis has been exploited to

promote vinylogous nucleophilicity within addition reactionmanifolds. We believe this reactivity may be further extendedto a variety of vinylogous donors and acceptors as well as tonucleophilic substitution reactions.

Methods and MaterialsAll the vinylogous adducts were fully characterized: Structural proofs andspectral data for all compounds are provided in the SI Appendix.

All the reactions were carried out in undistilled solvent without any pre-cautions to exclude moisture. To a solution of 9-amino(9-deoxy)epicinchonaalkaloids A–C (0.08 mmol) in 0.8 mL of toluene, ortho-fluorobenzoic acid(0.16 mmol, 22.4 mg) was added at room temperature under stirring.After 10 min, the reaction was started with the addition of β-substituted cy-clohexenone derivative 1 (0.8 mmol, 2.0 equiv) immediately followed by thenitroalkene 2 (0.4 mmol, 1.0 equiv), and the mixture was allowed to reach 40°C. Stirring was continued until complete conversion of the starting material(24–48 h, checked by thin layer chromatography). The reaction mixture wasthen directly purified by flash column chromatography (SiO2, 20–30% ethylacetate in hexane) to yield the vinylogous adducts 3 or 6.

ACKNOWLEDGMENTS.We thank F. Pesciaioli for fruitful discussions during theearly stage of the research project. G. Bergonzini is gratefully acknowledgedfor her experimental support. This work was supported by Bologna Univer-sity, the Catalan Institution for Research and Advanced Studies, and theInstitute of Chemical Research of Catalonia Foundation.

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