200,000+ products from a single source!
sales@angenechem.com
Home > Asymmetric Total Synthesis and Biological Evaluation of (+)-Cycloclavine
Asymmetric Total Synthesis and Biological Evaluation of (+)-Cycloclavine
Stephanie R. McCabe Peter Wipf
While the unique properties of naturally occurring compounds have always fascinated researchers from all branches of Science, the total synthesis of alkaloids currently experiences a remarkable renaissance, motivated by the complex architectures, diverse functionalities, and pro- found biological and cultural impact of this large family of natural products.1 Indole alkaloids, in particular, are attracting significant attention in Chemistry and Medicine.2 With the goal to explore both innovative synthetic strate- gies and new biological applications, we have recently es- tablished a program in the total synthesis of ergot alkaloids of the clavine and lysergic acid subclasses (Figure 1).
In 1969, A. Hofmann and co-workers at Sandoz in Basel, Switzerland, reported the isolation of a novel, cyclo- propane-containing ergot alkaloid, (+)-cycloclavine, from the seeds of the morning glory Ipomoea hildebrandtii VAT- KE, collected in Nairobi, Kenya.4 After a latency of almost 40 years, cycloclavine has now become a popular target for or- ganic synthesis, and a number of groups have reported innovative synthetic approaches. In pioneering studies from the group of Szántay, 4-bromo-Uhle’s ketone was subjected to an alkylation and intramolecular aldol reaction with 3- methylaminopropanoate, and the first synthesis of (±)-cy- cloclavine was completed in 2008 by a cyclopropanation of a tetrasubstituted alkene with CH2N2 (Scheme 1).5 Motivat- ed by our interests in the synthetic and medicinal chemis- try of indoles,6 the structurally unique indoline-indole scaf- fold of cycloclavine served as our first ergot alkaloid target. In 2011, we developed an intramolecular SN2-displacement and furan Diels–Alder reaction for the formation of the fused pentacyclic ring system and the synthesis of (±)-5- epi-cycloclavine.7 Subsequently, we modified this approach and used an intramolecular Diels–Alder reaction to a strain- activated methylenecyclopropane for the construction of the indoline segment and the total synthesis of (±)-cyclo- clavine (Scheme 1).
In 2014, Brewer and co-workers used a fragmentation and an azomethine ylide 1,3-dipolar cycloaddition to con- struct racemic cycloclavine (Scheme 2).8 Cao’s group developed two formal syntheses of racemic cycloclavine and a formal synthesis of (+)-cycloclavine starting from substitut- ed indoles and intersecting with the late stage alkene in Szántay’s synthesis.
Another formal synthesis of (±)-cycloclavine that con- verged with Szántay’s approach was accomplished by Netz and Opatz in 2016, utilizing a -alkylation of a pyrrolinone followed by a Heck coupling.
The first asymmetric synthesis of (–)-cycloclavine, the enantiomer of the natural alkaloid, was accomplished by our group in 2017.11 Key features of this synthesis were a catalytic asymmetric cyclopropanation of allene, an intra- molecular Diels–Alder reaction to methylenecyclopropane (IMDAMC), and an intramolecular Diels–Alder reaction to furan (IMDAF). Subsequently, a formal synthesis of both en- antiomers of cycloclavine was realized by Bisai and co- workers based on a D- or L-proline catalyzed -aminoxyl- ation and a Heck coupling (Scheme 2).12 Most recently, Dong and co-workers developed a benzyne cycloaddi- tion/alkene carboacylation route to both (–)-5-epi-cyclo- clavine and (–)-cycloclavine, utilizing a ring-enlargement of a benzocyclobutenone intermediate as a key reaction.13 The impressive publication surge and the diverse strategies of these synthetic approaches illustrate the high level of cur- rent interest in architecturally novel alkaloid natural prod- ucts. We now report the details of the first enantioselective total synthesis of (+)-cycloclavine.
Our retrosynthetic analysis is summarized in Scheme 3. In analogy to our route to (–)-cycloclavine,11 we selected an asymmetric rhodium-catalyzed cyclopropanation of allene with a diazopropanoate active ester, followed by an amino- lysis with 4-(methylamino)but-3-en-2-one, for the assem- bly of the key precursor for the IMDAMC reaction. After in- stalling the enone in the six-membered ring by a Diao– Stahl ketone dehydrogenation,14 the thermally removable Tempoc group for amine protection15 would be used to sta- bilize an aminomethyllithium reagent and favor enone 1,2- addition versus lactam ring opening. The final indole ring fusion was envisioned to be accomplished by the IMDAF cycloaddition.
For the realization of this retrosynthetic plan, two build- ing blocks and a chiral ligand needed to be prepared and optimized at the onset of the synthesis. The condensation of pyruvic acid (2) with tosyl hydrazide (1) under acidic conditions provided hydrazone 3 in 93% yield (Scheme 4). Treatment with oxalyl chloride and esterification of the resulting acid chloride with pentafluorophenol (PfOH) deliv- ered an active ester intermediate suitable for rapid segment assembly. Base-mediated diazo formation produced the first building block 4 in 21% yield.
Next, we focused on the selection of an appropriate transition metal catalyst and chiral ligand for the asymmet- ric allene cyclopropanation step. Diazopropanoates 5 are challenging reagents for use in metal-mediated cyclopro- panations because of the propensity of the metal carbenoid 6 to undergo competing -hydride migration to form an acrylic ester 7 (Scheme 5). In the past decade, several meth- ods have emerged that address this limitation. Fox et al. found that dirhodium complexes with sterically hindered carboxylate ligands in conjunction with low reaction tem- peratures effectively promoted intermolecular cyclopro- panations over the competing -H migration pathway.
Among the enantioselective variants, bulky carboxyl- ates derived from L-tert-leucine, such as Rh2(S-PTTL)4 (8), were found to be particularly effective. More recently, Hashimoto et al. showed that the substrate scope could be further expanded when the dirhodium complex Rh2(S-TBPTTL)4 (9) was used as the catalyst (Figure 2).18 We also prepared the enantiomer of 9, Rh2(R-TBPTTL)4(10), from D-tert-leucine [(R)-18] and anhydride 17 in toluene at reflux, followed by a ligand exchange reaction with Rh2(OAc)4 in a chlorobenzene/MeCN mixture at 130 °C (Scheme 6). Furthermore, we reasoned that the 4,7-diphe- nyl substitution pattern on the phthalimide ring of the nov- el, sterically demanding dirhodium catalyst 11 would im- pose even greater steric discrimination than the corre- sponding bromide substituents in 9 and 10, hopefully leading to greater differentiation between the enantiotopic faces of the allene. This catalyst was prepared in a Diels– Alder reaction of diphenylbutadiene (19) and maleic anhy- dride (20), followed by DDQ oxidation to afford anhydride 21, which was reacted with (S)-18 in the presence of tri- ethylamine to give 22 and subjected to a ligand exchange reaction to yield 11 (Scheme 7).
For further comparisons of ligand chemotypes, we de- cided to include an evaluation of the known dirhodium cat- alysts 12–14 (Figure 2).19 The ruthenium(II) complex 15 was added to this list because 15 was highly effective in re- lated asymmetric cyclopropanations of mono- and disubstituted allenes with succinimidyl diazoacetate.20 Finally, the (salen)cobalt(II) catalyst 16 was also screened since Katsuki et al. showed that it was an excellent catalyst for the enantioselective cyclopropanation of styrenes with -alkyl- diazoacetates.21
The results of the cyclopropanation of allene (23) with pentafluorophenyl diazopropanoate (4) to give methy- lenecyclopropane 24 in the presence of the chiral catalysts 8–16 are summarized in Table 1. Rh(II)-Catalysts with steri- cally hindered amino acid ligands but lacking phthalimide substituents, such as 8 and 12, provided a low e.r. of ap- proximately 7:3 (Table 1, entries 1 and 5). Hashimoto’s tetrabromophthaloyl tert-leucine dirhodium catalyst 9 re- sulted in a notable improvement, giving the cyclopropane (R)-24 in a high yield with an e.r. of 87:13 (entry 2). As ex- pected, the (R)-tert-leucine derived 10 gave the enantio- meric product (S)-24 in identical yield and e.r. (entry 3). Disappointingly, however, virtually no enantioinduction was observed when allene was reacted with 4 in the pres- ence of the sterically more demanding dirhodium catalyst 11 (product e.r. = 55:45, entry 4). With this catalyst, no re- action occurred at –78 °C and the mixture had to be warmed to –40 °C before conversion was observed. The chi- ral cyclopropane catalyst 13 delivered the desired product 24 in moderate yields and with poor enantioinduction (en- try 6). Davies’ proline-based catalyst 14 also provided only a moderate yield of 61%, and negligible asymmetric induc- tion (entry 7). The reaction of Ru(II)-catalyst 15 did not de- liver any of the desired product. Instead, upon warming the reaction mixture from –78 °C to room temperature, full conversion into the undesired -H migration product 25 was observed (entry 8). Similarly, the (salen)cobalt(II) cata- lyst 16 showed no catalytic activity even at room tempera- ture (entry 9).
In our synthetic route, the cyclopropanation of allene(23) with diazopropanoate 4 in the presence of 1 mol% of the dirhodium catalyst Rh2(R-TBPTTL)4 (10) provided the enantiomerically enriched methylenecyclopropane (S)-24 on multi-gram scale. Ester aminolysis with the lithium salt of 26 gave the vinylogous imide 27 in 80% yield and 87:13 e.r. (Scheme 8). Deprotonation of 27 with NaHMDS in THF at –78 to –50 °C formed the corresponding sodium enolate, which was trapped with TBSCl to give the silyl enol ether intermediate 28. When heated at 95 °C in THF in the microwave reactor, this diene underwent the intramolecular strain-promoted Diels–Alder (IMDAMC) reaction, and TBAF cleavage of the product silyl enol ether formed the desired trans-adduct 30 in 66% yield along with the cis-adduct 29 in 15% yield. These two diastereomers were readily separated chromatographically. Dehydrogenation of 30 under modified Diao–Stahl conditions14 with Pd(TFA)2 and DMSO in AcOH under an atmosphere of oxygen at 55 °C gave the corresponding enone 31 in 66% yield as a single regioisomer. Interestingly, dehydrogenation of the epimer 29 under these conditions provided the opposite enone regioisomer 32 stereospecifically in 56% yield. This complete switch in regioselectivity for the cis- and trans-diastereomers 29 and 30 parallels results obtained for the enolization of cis- and trans-2-decalones.22 The configuration at the decalin ring junction governs the regioselectivity of enolization process due to torsional strain effects. The torsional strain that was proposed to govern the regiochemistry of enolization in cis- and trans-decalones has also been investigated in greater detail in relevant cis- and trans-octalins.
Enone 31 was obtained as a crystalline solid, and its en- antiomeric ratio could be further enriched by recrystalliza- tion from 87:13 to yield product with >99% e.r. Chiral SFC analysis was used to evaluate each batch for enantiomeric purity.
For the completion of the total synthesis, stannane 34 was treated with n-BuLi at low temperature and converted into the corresponding lithium carbanion (Scheme 9). Addi- tion of enone 31 to the reaction mixture delivered two dia- stereomeric allylic alcohols, which could be separated by chromatography on silica gel to give -alcohol -35 and alcohol -35 in 41% and 32% yield, respectively. The intra- molecular Diels–Alder (IMDAF) reaction of -35 (bearing a pseudo-equatorial hydroxy group) at 135 °C in a sealed tube was followed by spontaneous aromatization and cleav- age of the Tempoc protecting group under the reaction con- ditions to form indole 36. The stereoisomeric alcohol -35 was inert under these conditions, quite likely due to steric strain in the transition state that requires a pseudo-axial position of the substituent bearing the furan ring. Finally, lactam reduction led to (+)-cycloclavine in 34% yield over two steps from -35. Overall, the synthesis of the natural enantiomer was accomplished in 8 steps and 4% yield. The specific rotation of the synthetic material was determined to be +61.4 (c 0.2, CHCl3), which was consistent with the literature value +63 (c 1, CHCl3).4 Mass spectra, IR, 1H, and 13 C NMR data were also consistent with the previously re- ported data for the natural product as well as its enantiomer.
In contrast to the vast literature on lysergic acid deriva- tives, relatively little is known about the pharmaceutical potential of clavine ergot alkaloids.2b Lysergic acid deriva- tives, most notably lysergic acid diethylamide (LSD) have significant hallucinogenic properties that can interfere with their therapeutic potential. Most of these effects are thought to be mediated through agonist action at the 5-hy- droxytryptamine receptor 2A, 5-HT2A. In the tailwind of the rapidly expanding medical uses of cannabinoids, the mush In contrast to the vast literature on lysergic acid deriva- tives, relatively little is known about the pharmaceutical potential of clavine ergot alkaloids.2b Lysergic acid deriva- tives, most notably lysergic acid diethylamide (LSD) have significant hallucinogenic properties that can interfere with their therapeutic potential. Most of these effects are thought to be mediated through agonist action at the 5-hy- droxytryptamine receptor 2A, 5-HT2A. In the tailwind of the rapidly expanding medical uses of cannabinoids, the mush room metabolite psilocybin, and even LSD, are now moving to the forefront of clinical research on the management of mental health, anxiety, neurodegeneration, and substance- use disorders.24 It would appear that more fundamental re- search on efficacy, tolerability, and safety of serotonergic psychedelics is highly warranted.
It is well known that binding of lysergic acid derivatives to brain membrane receptors is stereospecific, since L-LSD, the psychotropically inactive enantiomer of LSD, is ca. 1000 times weaker as a brain membrane receptor radioligand displacing agent,25 and L-LSD as well as the other diastereo- mers, D-iso-lysergic acid diethylamide (iso-LSD) and L-iso- lysergic acid diethylamide (L-iso-LSD), show no psychic ef- fects in humans up to a dose of 0.5 mg, which corresponds to a 20-fold increase over a still distinctly active D-LSD dose.26 Having gained ready synthetic access to both natural (+)-cycloclavine and its unnatural enantiomer (–)-cyclo- clavine,11 we were therefore interested to determine the re- ceptor profiles of both compounds, and compare them to other serotonergic agents.
For an initial survey, we selected 13 pertinent CNS re- ceptors and profiled both enantiomers at 10 M concentra- tion (Table 2). Compound binding was calculated as a per- cent inhibition of a radioactively labeled ligand specific for each target. As a group, the cycloclavines were more selec- tive in this receptor panel than D-LSD,27,28 the bioactive LSD stereoisomer. D-LSD was active at the adrenergic 1 and his- tamine H1 receptors (Table 2, entries 1 and 6), whereas both cycloclavines were moderately active at the opiate recep- tor (entry 10). Neither ergot chemotype showed significant activity at GABAA, muscarinic M2 and M5, and nicotinic ace- tyl-choline 42 receptors (entries 5, 7, 8, and 9). We were unable to find LSD data on orexin OX1, but cycloclavine did not perturb radioligand binding at this site at a 10 M con- centration (entry 11).
Significant differences between (+)- and (–)-cyclo- clavine revealed themselves in the dopamine D1, D2L, and D3 monoamine receptor family (Table 2, entries 2–4). In close analogy to D-LSD, natural (+)-cycloclavine maintained strong affinity to these receptors, which stimulate cognitive and motor functions. (–)-Cycloclavine showed compara- tively moderate activity at the dopamine D3 receptor at 10 M concentration, but fell below the threshold of 50% inhi- bition at 1 M, whereas (+)-cycloclavine still maintained significant binding at this concentration (entry 4). A less prominent but still distinctive stereospecificity was ob- served at the serotonin 5-HT1A and 5-HT2A receptors (en- tries 12 and 13). Natural (+)-cycloclavine had very potent binding properties at both 10 and 1 M, whereas (–)-cyclo- clavine tailed off at 1 M. Serotonin receptors regulate a plethora of behavioral responses, from aggression, anxiety, appetite, to learning, memory, sleep, and even aging.29 D- LSD is one of the most potent agonists at 5-HTA, and the af- finity at the 5-HTA2 and possibly the 5-HT2C receptors ver- sus the 5-HT1A receptor correlates with the mental effects of psychedelics in humans.30 In view of this interesting ste- reospecificity, and the significance of 5-HT receptors to hu- man behavior, we decided to pursue additional studies on 5-HT subtypes (Table 3).
The purpose of our second generation functional assays on human 5-HT receptors was to determine effective con- centrations EC50 or inhibitory constants (Ki) for (+)-cyclo- clavine and (–)-cycloclavine. Cellular agonist effects were calculated as a percentage of a control response to a validat- ed reference for each target, and cellular antagonist effect was calculated as percent inhibition of a validated control agonist response for each target. In addition to D-LSD, we selected N,N-dimethyltryptamine (DMT) and psilocin as two relevant reference compounds.31,32 DMT is the only known endogenous N,N-dimethylated trace amine in mam- mals, and a prominent component in the sacramental tea ayahuasca.33 Its psychopharmacology has recently been compared to so-called ‘near-death experiences’.34 Psilocin is the pharmacologically active agent after ingestion of the prodrug psilocybin present in some species of psychedelic mushrooms. Psilocybin is currently clinically investigated as a treatment for anxiety and depression in cancer care, as well as for enhancement of cognitive flexibility and creativ- ity.
As suggested by the preliminary assays, (+)-cycloclavine provided considerably more potent at the 5-HT1A receptor than (–)-cycloclavine with an activation potency EC50 = 0.14M versus ~5 M for (–)-cycloclavine (Table 3, entry 1). Both stereoisomers are poor activators at 5-HT2A, suggest- ing that hallucinogenic or strongly euphoric effects in hu- mans might be limited in comparison to D-LSD, even though (+)-cycloclavine displays its most potent activation potential EC50 = 16 nM at 5-HT2C, a receptor that is thought to contribute to the observed mental effects of psychedelic drugs (entries 2 and 4). With the exception of DMT, which has only moderate potency, none of the tested agents acti- vated 5-HT2B, a 5-HT receptor subtype that has been associ- ated with cardiotoxicity. Overall, the 5-HT profile of (+)-cy- cloclavine closely mirrors that of psilocin, and to a lesser ex- tent, that of DMT. It is substantially different from D-LSD, a property that we believe bodes well for future therapeutic investigations of this compound class.
The unusual activity on the opioid receptor, and the relative similarity to psilocin and DMT in the 5-HT panel in- spired us to also evaluate the activity of cycloclavines in the sigma-1 assay, a receptor that was originally mischaracter- ized as an opioid receptor and has now been implicated in neuroinflammation and neuroprotection.36 DMT was iden- tified as an endogenous sigma-1 receptor regulator.33,37 Sur- prisingly, while (+)-cycloclavine was inactive, the unnatural (–)-cycloclavine was determined to have a Ki = 8.3 M for the inhibition of the binding of the radiolabeled agonist hal- operidol to sigma-1, and therefore found to be very similar to DMT (Ki = 5.2 M) (Table 3, entry 5). To the best of our knowledge, this is the first time that stereospecific binding of ergot alkaloids to a sigma receptor has been observed, and, accordingly, it is feasible to consider (–)-cycloclavine as a potential lead structure for sigma receptor modulator design.
In conclusion, we have successfully completed a total synthesis of natural (+)-cycloclavine, featuring an optimiza- tion of the catalyst for the asymmetric cyclopropanation of allene with an active ester diazopropanoate, a regiospecific Pd-catalyzed ketone dehydrogenation to the enone, and two intramolecular Diels–Alder reactions for indole/indo- line annulations. Furthermore, we have characterized the binding effects of (+)- and (–)-cycloclavine against 16 CNS receptors, and discovered significant stereospecificity prop- erties. (+)-Cycloclavine has at least 10-fold higher potency at the serotonin 5-HT2C receptor than at any of the other tested receptors, making it one of the most selective trypt- amines discovered to date. Furthermore, the receptor sub- type profile of (+)-cycloclavine resembles that of the clini- cally validated mushroom metabolite psilocin more closely than the related psychedelics LSD and DMT. Finally, we de- termined that the unnatural (–)-cycloclavine has consider- ably lower affinities at all 5-HT receptors than (+)-cyclo- clavine, but is quite active at the sigma-1 receptor, a proper- ty that it shares with the endogenous sigma-1 ligand DMT. We suggest that these results, in combination with the ex- cellent synthetic tractability of the cycloclavine scaffold, encourage future research on the medicinal chemistry of clavine alkaloids.
ANGENE is pledged to providing quality specialty chemicals for use in research and development and commercial manufacturing:
© 2019 Angene International Limited. All rights Reserved.