11H-Benzo[4,5]imidazo[1,2-a]indol-11-one as a New Precursor of Azomethine Ylides: 1,3-Dipolar Cycloaddition Reactions with Cyclopropenes and Maleimides

The possibility of generating azomethine ylides from 11H-benzo[4,5]imidazo[1,2-a]indol-11-one and amino acids is shown for the first time. Based on the cycloaddition reactions of these azomethine ylides with cyclopropenes and maleimides, cyclopropa[a]pyrrolizines, 3-azabicyclo[3.1.0]hexanes, and pyrrolo[3,4-a]pyrrolizines spiro-fused with a benzo[4,5]imidazo[1,2-a]indole fragment were synthesized. Spirocyclic compounds were obtained in moderate to good yields, albeit with poor diastereoselectivity. Density functional theory calculations were performed to obtain an insight into the mechanism of the 1,3-dipolar cycloaddition of 11H-benzo[4,5]imidazo[1,2-a]indol-11-one-derived azomethine ylides to cyclopropenes. The cytotoxic activity of some of the obtained cycloadducts against the human erythroleukemia (K562) cell line was evaluated in vitro by MTS-assay.


Introduction
1,3-Dipolar cycloaddition (1,3-DC) is one of the most versatile methodologies for constructing five-membered heterocycles [1,2]. [3 + 2] Cycloaddition can take place with the participation of various 1,3-dipoles and dipolarophiles; however, these reactions have several features in common: (1) the dipole has a charge, (2) the resulting cycloadduct is uncharged, and (3) the HOMO of the 1,3-dipole reacts with the LUMO of the dipolarophile (a HOMO-controlled reaction). Among the 1,3-dipoles, azomethine ylides (belonging to allyl-type N-centered dipoles) are among the most popular and frequently used in 1,3-dipolar cycloaddition reactions [3,4]. One of the most well-known methods for the preparation of azomethine ylides is the reaction of amines with aldehydes, followed by deprotonation of the iminium cation or a prototropic shift of the imine [5,6]. Amino acids are capable of reacting with aldehydes or ketones to generate intermediate oxazolidinones, which undergo decarboxylation, resulting in the formation of azomethine ylides [7,8]. The other commonly used approaches to generate azomethine ylides rely on thermal ring-opening reactions involving aziridines or 4-oxazolines [9,10]. These and other methods continue to be widely used in synthetic organic chemistry. Reactions of azomethine ylides with alkenes make it possible to obtain highly substituted pyrrolidines containing up to four stereocenters [11,12]. The pyrrolidine ring, isolated or fused, can be attributed to a fundamental structural fragment that is part of a large number of natural and other methods continue to be widely used in synthetic organic chemistry. Reactions of azomethine ylides with alkenes make it possible to obtain highly substituted pyrrolidines containing up to four stereocenters [11,12]. The pyrrolidine ring, isolated or fused, can be attributed to a fundamental structural fragment that is part of a large number of natural and synthetic biologically-significant compounds [13]. The analysis of 164 U.S. FDA-approved small-molecule drugs from 2015 to June 2020 revealed that 22 (13%) of them contain a pyrrolidine moiety [14]. At the same time, 17 (8.5%) out of 200 small molecular drugs with the largest retail sales in 2020 contained the pyrrolidine cycle [15]. Nevertheless, despite many achievements in the study of 1,3-dipolar cycloaddition reactions, the search for new carbonyl substrates for the generation of azomethine ylides, which can lead to the construction of new heterocyclic systems containing valuable pharmacophore fragments, is highly desirable. Of the pharmacophore fragments that can be obtained through the [3 + 2] cycloaddition reactions of azomethine ylides, in our opinion, derivatives of 3-azabicyclo[3.1.0]hexane and pyrrolizine deserve special attention ( Figure 1). 3-Azabicyclo[3.1.0]hexanes can be characterized as an important class of heterocyclic compounds with diverse pharmacological and biological activities. For example, 3-azabicyclo[3.1.0]hexanes exhibit anti-neuroinflammatory (monoacylglycerol lipase inhibitor, (I) [16], anti-neurodegenerative (dual leucine zipper kinase inhibitor, (II) [17] and antiviral (SARS-CoV-2 main protease inhibitor, (III) [18,19] effects. It is well known that pyrrolizine is the main structural moiety in many organic compounds exhibiting various biological activities, including anticoagulant (serine protease thrombin inhibitor, (IV) [20], anti-cancer (indicine N-oxide, (V), is an antitumor agent for pediatric cancer and solid tumors research) 14 and anti-HIV activities (7,7a-diepialexine, (VI) [21] ( In previous studies bys our research group, cyclopropenes were widely used as dipolarophiles in 1,3-dipolar cycloaddition reactions with azomethine ylides generated from the corresponding ketones and -amino acids (Scheme 1). We used derivatives of isatin [22], alloxan [23], ninhydrin [24][25][26][27], 11H-indeno [1,2-b]quinoxalin-11-one [28], and tryptanthrin [29] as the ketone components. The products of these reactions were pharmacologically interesting spiro-fused 3-azabicyclo[3.1.0]hexanes and pyrrolizines, some of which were identified as exhibiting in vitro antitumor activity [30,31]. In previous studies bys our research group, cyclopropenes were widely used as dipolarophiles in 1,3-dipolar cycloaddition reactions with azomethine ylides generated from the corresponding ketones and α-amino acids (Scheme 1). We used derivatives of isatin [22], alloxan [23], ninhydrin [24][25][26][27], 11H-indeno [1,2-b]quinoxalin-11-one [28], and tryptanthrin [29] as the ketone components. The products of these reactions were pharmacologically interesting spiro-fused 3-azabicyclo[3.1.0]hexanes and pyrrolizines, some of which were identified as exhibiting in vitro antitumor activity [30,31].
As we noted above, the search for new substrates for the generation of azomethine ylides is an urgent task, since it allows introducing new unique poly/spirocyclic systems into the arsenal of heterocyclic chemistry, which may be of interest for pharmacology. With our continued research interest in 1,3-dipolar cycloaddition reactions, herein we present the first example of the generation of azomethine ylides from the tetracyclic ketone 11H-benzo [4,5]imidazo [1,2-a]indol-11-one and an α-amino acids, and also its [3 + 2] cycloaddition with cyclopropenes and maleimides (Scheme 1). It is known that indole-fused heterocycles, such as benzoimidazo [1,2-a]indole derivatives, are structural fragments of numerous natural products and pharmacologically active compounds [32][33][34][35][36]. Previously, Lee and coworkers [37,38] described the functionalization of 11H-benzo [4,5]imidazo [1,2a]indol-11-one, which resulted in compounds of interest for the creation of phosphorescent organic light-emitting devices.
As we noted above, the search for new substrates for the generation of azomethine ylides is an urgent task, since it allows introducing new unique poly/spirocyclic systems into the arsenal of heterocyclic chemistry, which may be of interest for pharmacology. With our continued research interest in 1,3-dipolar cycloaddition reactions, herein we present the first example of the generation of azomethine ylides from the tetracyclic ketone 11H-benzo [4,5]imidazo [1,2-a]indol-11-one and an -amino acids, and also its [3 + 2] cycloaddition with cyclopropenes and maleimides (Scheme 1). It is known that indole-fused heterocycles, such as benzoimidazo [1,2-a]indole derivatives, are structural fragments of numerous natural products and pharmacologically active compounds [32][33][34][35][36]. Previously, Lee and coworkers [37,38] described the functionalization of 11H-benzo [4,5]imidazo [1,2-a]indol-11-one, which resulted in compounds of interest for the creation of phosphorescent organic light-emitting devices.

Synthesis
In initial studies, the possibility of 1,3-dipolar cycloaddition was investigated by performing a model three component reaction of cyclopropene 1a, 11H-benzo [4,5]imidazo [1,2a]indol-11-one (2), and L-proline (3a) in various solvents (Table 1). In methanol or ethanol, only trace amounts of cycloadduct 4a/5a could be detected (Table 1, entries 1 and 2); when the reaction was carried out in THF or acetonitrile, the desired product 4a/5a was obtained in 21% or 52%, respectively, after 18 h (Table 1, entries 3 and 4). At the same time, when carrying out the reaction in low-polarity dioxane, it was possible to increase the yield of the target product to 64%; however, in this case, as in all others, a low level of diastereoselectivity was observed (dr 2:1) ( Table 1, entry 5). The use of non-polar solvents such as benzene or toluene did not increase the yield (Table 1, entries 6 and 7). Non-polar solvents such as benzene or toluene did not facilitate an increase of the yield (Table 1, entries 6 and 7). Through the experiments above, we finally confirmed the optimal reaction conditions: 1 (0.3 mmol), 2 (0.3 mmol), 3 (0.6 mmol), and 1,4-dioxane (4 mL) in a sealed tube at 100 • C for 18 h under a nitrogen atmosphere.
To explore the scope of the reaction, a range of cyclopropene dipolarophiles 1a-h was tested in the reaction with azomethine ylide generated from ketone 2 and L-proline (3a) under optimized conditions. The results are summarized in Table 2. yield (Table 1, entries 6 and 7). Non-polar solvents such as benzene or tolu facilitate an increase of the yield (Table 1, entries 6 and 7). Through the e above, we finally confirmed the optimal reaction conditions: 1 (0.3 mmol), 2 ( (0.6 mmol), and 1,4-dioxane (4 mL) in a sealed tube at 100 °C for 18 h unde atmosphere. To explore the scope of the reaction, a range of cyclopropene dipolaro was tested in the reaction with azomethine ylide generated from ketone 2 an (3a) under optimized conditions. The results are summarized in Table 2.   In addition to product 4a, various cycloadducts can be prepared as mixtures of d stereomers with good conversion and low dr ( Table 2). In most cases, the diastereome are chromatographically inseparable; however, the major endo-diastereomer 4 can isolated by crystallization of the mixture from methanol ( Table 2, 4a-c,e-h). Only in t case of cycloadduct 4d/5d derived from trimethylsilylcyclopropene 1d were both d stereomers 4d and 5d successfully separated by preparative TLC. It is important to no that cyclopropenes with both electron-rich and electron-deficient substituents R 1 at t  In addition to product 4a, various cycloadducts can be prepared as mixtures of diastereomers with good conversion and low dr ( Table 2). In most cases, the diastereomers are chromatographically inseparable; however, the major endo-diastereomer 4 can be isolated by crystallization of the mixture from methanol ( Table 2, 4a-c,e-h). Only in the case of cycloadduct 4d/5d derived from trimethylsilylcyclopropene 1d were both diastereomers 4d and 5d successfully separated by preparative TLC. It is important to note that cyclopropenes with both electron-rich and electron-deficient substituents R 1 at the C3 position turned out to be reactive towards this azomethine ylide. The corresponding cycloadducts 4a-h were obtained as mixtures of two diastereomers in overall yields of 58- In addition to product 4a, various cycloadducts can be prepared as mixtures of diastereomers with good conversion and low dr ( Table 2). In most cases, the diastereomers are chromatographically inseparable; however, the major endo-diastereomer 4 can be isolated by crystallization of the mixture from methanol ( Table 2, 4a-c,e-h). Only in the case of cycloadduct 4d/5d derived from trimethylsilylcyclopropene 1d were both diastereomers 4d and 5d successfully separated by preparative TLC. It is important to note that cyclopropenes with both electron-rich and electron-deficient substituents R 1 at the C3 position turned out to be reactive towards this azomethine ylide. The corresponding cycloadducts 4a-h were obtained as mixtures of two diastereomers in overall yields of 58-83% (Table 2). No significant effect of the R 1 substituents on the course of the reactions was observed. The structures of major diastereomers 4 were unambiguously confirmed by NMR spectroscopy and single-crystal X-ray analysis of 4f ( Figure 2). The stereochemistry of the minor diastereomer 5d was determined using 2D NMR spectroscopy (see the Supporting Information, Figures S53-S59). As can be seen, the diastereomers 4 and 5 differ from each other in the configuration of the spiro atom. Attempts to carry out this reaction with other secondary α-amino acids such as sarcosine, azetidine-2-carboxylic acid, pipecolic acid, and thiazolidine-4-carboxylic acid were unsuccessful. It is noteworthy that only cyclopropenes containing phenyl substituents at the double bond can be used in this reaction. Unsubstituted and methyl-substituted cyclopropenes proved to be ineffective as dipolarophiles in this reaction. The trimethylsilyl group of 4d was readily removed by K2CO3 in methanol and ether to obtain cyclopropylacetylene 4i in a 92% yield (Scheme 2). The trimethylsilyl group of 4d was readily removed by K 2 CO 3 in methanol and ether to obtain cyclopropylacetylene 4i in a 92% yield (Scheme 2). Upon further study of the substrate scope, we focused on the α-amino acid component (Table 3). First, we performed a reaction between cyclopropene 1a and azomethine ylide generated in situ from ketone 2 and 2-aminobutanoic acid (3b). As a result, spiro[3-azabicyclo[3.1.0]hexane] 6a was obtained as a chromatographically inseparable mixture of two diastereomers in 46% yield with low dr (2.6:1). The major endo-diastereomer can be obtained in pure form by crystallization from methanol. The reaction between cyclopropene 1a, ketone 2, and different alkyl-substituted amino acids (DL-norvaline (3c), L-methionine (3d), DL-norleucine (3e), and L-leucine (3f)) proceeded smoothly under the optimal reaction conditions and produced the expected spiro cycloadducts 6b-e in moderate isolated yields (42-53%) ( Table 3). As in the previous case, these products are formed as a mixture of diastereomers, from which the main component can be isolated by crystallization from methanol. Subsequently, the cyclopropene scope of the 1,3-dipolar cycloaddition was investigated, employing ketone 2 and L-leucine (3f) as precursors of the azomethine ylide. It should be noted that cyclopropenes 1b-d,f,h give the corresponding products 6f-j in moderate yields, regardless of the electronic effect of the substituent at C3 (Table 3). Summarizing the data given in Table 3, it should be noted that the nature of the substituent at the cyclopropene ring does not Upon further study of the substrate scope, we focused on the α-amino acid component (Table 3). First, we performed a reaction between cyclopropene 1a and azomethine ylide generated in situ from ketone 2 and 2-aminobutanoic acid (3b). As a result, spiro[3azabicyclo[3.1.0]hexane] 6a was obtained as a chromatographically inseparable mixture of two diastereomers in 46% yield with low dr (2.6:1). The major endo-diastereomer can be obtained in pure form by crystallization from methanol. The reaction between cyclopropene 1a, ketone 2, and different alkyl-substituted amino acids (DL-norvaline (3c), L-methionine (3d), DL-norleucine (3e), and L-leucine (3f)) proceeded smoothly under the optimal reaction conditions and produced the expected spiro cycloadducts 6b-e in moderate isolated yields (42-53%) ( Table 3). As in the previous case, these products are formed as a mixture of diastereomers, from which the main component can be isolated by crystallization from methanol. Subsequently, the cyclopropene scope of the 1,3-dipolar cycloaddition was investigated, employing ketone 2 and L-leucine (3f) as precursors of the azomethine ylide.
It should be noted that cyclopropenes 1b-d,f,h give the corresponding products 6f-j in moderate yields, regardless of the electronic effect of the substituent at C3 (Table 3). Summarizing the data given in Table 3, it should be noted that the nature of the substituent at the cyclopropene ring does not significantly affect the yield of the target spiro adducts nor the stereochemistry of cycloaddition. The stereochemistry of the major diastereomer is identical to the stereochemistry of the cycloadducts obtained from cyclopropenes and azomethine ylides based on isatin, 11H-indeno[1,2-b]quinoxalin-11-one, and tryptanthrin (previously described in a series studies) [22,28,29]. In the case of other primary α-amino acids such as glycine, alanine, valine, 2-aminocaprylic acid, phenylglycine, phenylalanine, and tryptophan, the corresponding cycloadducts could not be obtained.  As shown above, cyclopropenes can successfully act as dipolarophiles in reactions with azomethine ylides derived from ketones and amino acids, which allowed us to expand the structural diversity of dipoles capable of reacting with cyclopropenes. However, other dipolarophiles are also of interest for studying their reactivity towards new azomethine ylides. We turned our attention to the maleimides, classical dipolarophiles well-studied in many 1,3-dipolar cycloaddition reactions. Various maleimides 7 were tested in the reaction with azomethine ylide generated from 2 and 3a, and the results are summarized in Table 4. Optimization of the reaction conditions showed that the best yield of cycloadducts was achieved when the reactions were carried out in an acetonitrile medium. The three-component reaction involving parent maleimide (7a) proceeded in acetonitrile at 100 °C for 18 h with the formation of endo-product 8a in 78% yield with 1.5:1 diastereoselectivity ( Table 4). Reaction of ketone 2, L-proline (3a) and N-methyl maleimide (7b) afforded 8b in 69% yield with low diastereoselectivity (1.7:1 dr). Interestingly, N-phenyl maleimide (7c) resulted in an increase in diastereoselectivity (8c, 9:1 dr). The higher diastereoselectivity can be explained by the π-π interaction between the phenyl group of maleimide 7c and the aromatic moiety of azomethine ylide.  As shown above, cyclopropenes can successfully act as dipolarophiles in reactions with azomethine ylides derived from ketones and amino acids, which allowed us to expand the structural diversity of dipoles capable of reacting with cyclopropenes. However, other dipolarophiles are also of interest for studying their reactivity towards new azomethine ylides. We turned our attention to the maleimides, classical dipolarophiles well-studied in many 1,3-dipolar cycloaddition reactions. Various maleimides 7 were tested in the reaction with azomethine ylide generated from 2 and 3a, and the results are summarized in Table 4. Optimization of the reaction conditions showed that the best yield of cycloadducts was achieved when the reactions were carried out in an acetonitrile medium. The three-component reaction involving parent maleimide (7a) proceeded in acetonitrile at 100 °C for 18 h with the formation of endo-product 8a in 78% yield with 1.5:1 diastereoselectivity ( Table 4). Reaction of ketone 2, L-proline (3a) and N-methyl maleimide (7b) afforded 8b in 69% yield with low diastereoselectivity (1.7:1 dr). Interestingly, N-phenyl maleimide (7c) resulted in an increase in diastereoselectivity (8c, 9:1 dr). The higher diastereoselectivity can be explained by the π-π interaction between the phenyl group of maleimide 7c and the aromatic moiety of azomethine ylide. N-Alkylmaleimides 7d-f also reacted smoothly with the azomethine ylide generated from 2 and 3a to form the desired endo-products 8d-f in high yields (68−89%), albeit with a Reactions of 1 (0.3 mmol), 2 (0.3 mmol), and 3 (0.6 mmol) were carried out in 1,4-dioxane (4 mL) in a sealed tube at 100 • C for 18 h under a nitrogen atmosphere. b Isolated yield. c Crystallization of the diastereomeric mixture from methanol gave the major diastereomer 6 as a pure compound. d The dr values were determined by 1 H NMR of the crude mixture.
As shown above, cyclopropenes can successfully act as dipolarophiles in reactions with azomethine ylides derived from ketones and amino acids, which allowed us to expand the structural diversity of dipoles capable of reacting with cyclopropenes. However, other dipolarophiles are also of interest for studying their reactivity towards new azomethine ylides. We turned our attention to the maleimides, classical dipolarophiles well-studied in many 1,3-dipolar cycloaddition reactions. Various maleimides 7 were tested in the reaction with azomethine ylide generated from 2 and 3a, and the results are summarized in Table 4. Optimization of the reaction conditions showed that the best yield of cycloadducts was achieved when the reactions were carried out in an acetonitrile medium. The threecomponent reaction involving parent maleimide (7a) proceeded in acetonitrile at 100 • C for 18 h with the formation of endo-product 8a in 78% yield with 1.5:1 diastereoselectivity ( Table 4). Reaction of ketone 2, L-proline (3a) and N-methyl maleimide (7b) afforded 8b in 69% yield with low diastereoselectivity (1.7:1 dr). Interestingly, N-phenyl maleimide (7c) resulted in an increase in diastereoselectivity (8c, 9:1 dr). The higher diastereoselectivity can be explained by the π-π interaction between the phenyl group of maleimide 7c and the aromatic moiety of azomethine ylide. N-Alkylmaleimides 7d-f also reacted smoothly with the azomethine ylide generated from 2 and 3a to form the desired endo-products 8d-f in high yields (68−89%), albeit with low diastereoselectivity. The structure and relative configuration of 8f were unambiguously determined by X-ray diffraction analysis (Figure 3). In line with the structure of 8f, this relative configuration was assigned to other major cycloadducts 8. The minor diastereomer is an epimer and differs in its spiro atom configuration. this relative configuration was assigned to other major cycloadducts 8. The minor diastereomer is an epimer and differs in its spiro atom configuration. this relative configuration was assigned to other major cycloadducts 8. The stereomer is an epimer and differs in its spiro atom configuration.

Computational Study
Given that 11H-benzo [4,5]imidazo [1,2-a]indol-11-one (2) had not previously been studied as a precursor of azomethine ylides, we considered it necessary to carry out a density functional theory (DFT) computational study that would enable obtaining a comprehensive view of the reaction mechanisms of azomethine ylide formation and cycloaddition between 1,2-diphenyl-3-vinylcyclopropene (1c) and azomethine ylide generated from 2 ( Figure 4). Moreover, this calculation data may provide further evidence of the relative configuration of both resulting diastereomers 4c and 5c.

Biological Assay
The antiproliferative activity of some of the synthesized spiro-fused 11H-benzo [4,5]imidazo[1,2-a]indol-11-ones and cyclopropa [a]pyrrolysines 4 or 3-azabicyclo[3.1.0]hexanes 6 against human erythroleukemia (K562) cell line was evaluated in vitro using the standard MTS assay for 24 and 72 h. The results of these investigations are presented in Figure 5. It was found that spiroadducts with the N-isopropylcarbamoyl group at the cyclopropane moiety were more active, while replacement of this group with phenyl one led to a significant decrease in this activity. Thus, it was found that among the tested compounds, spiroadducts 4f and 6i demonstrated a significant activity, with IC 50 5 ± 1 and 12 ± 2 µg/mL, respectively. kcal/mol). The calculation data are completely consistent with experimental results. Actually, cycloadduct 4c, the precursor of which is the more thermodynamically stable ylide AY-1, predominates in the mixture of diastereomers 4c and 5c. In the meantime, minor diastereomer 5c results from the endo approach of ylide AY-2 to cyclopropene 1c. Probably, these findings hold true for other cycloaddition reactions between cyclopropenes 1 and azomethine ylide generated from 2 and 3a.

Biological Assay
The antiproliferative activity of some of the synthesized spiro-fused 11H-benzo [4,5]imidazo [1,2-a]indol-11-ones and cyclopropa [a]pyrrolysines 4 or 3-azabicyclo[3.1.0]hexanes 6 against human erythroleukemia (K562) cell line was evaluated in vitro using the standard MTS assay for 24 and 72 h. The results of these investigations are presented in Figure 5. It was found that spiroadducts with the N-isopropylcarbamoyl group at the cyclopropane moiety were more active, while replacement of this group with phenyl one led to a significant decrease in this activity. Thus, it was found that among the tested compounds, spiroadducts 4f and 6i demonstrated a significant activity, with IC50 5 ± 1 and 12 ± 2 μg/mL, respectively.
Yield  [53]. Thermal corrections to enthalpy and entropy values were evaluated at 298.15 K and 1.0 atm. All calculations were performed using the Gaussian 09 computational program package [54]. Data on energies (a.u.) and cartesian coordinates of stationary points for reactants, intermediates, products and the transition states (M062x/cc-pVDZ, PCM = 1,4-dioxane) are given at Supporting Information, (Table S5).

Cell Culture and Culturing Conditions
The human erythroleukemia (K-562) cell line was obtained from the cell repository "Vertebrate cell culture collection" (supported by the Ministry of Science and Higher Education of the Russian Federation, agreement № 075-15-2021-683, Institute of Cytology, Russian Academy of Sciences, Saint Petersburg, Russia). Cells were grown on RPMI medium (Hyclone, GE Healthcare Life Sciences, Logan, UT, USA) supplemented with 10% (v/v) fetal bovine serum (Hyclone, GE Healthcare Life Sciences, Logan, UT, USA) and gentamicin (Sigma-Aldrich, St. Louis, MO, USA) at 37 • C in a humidified atmosphere with 5% CO 2 .

Cell Proliferation Assay
To evaluate the in vitro toxicity of the compounds synthesized, cells were seeded into 96-well plates at a density of 5 × 10 3 cells per well. On the next day, the tested compounds were added to the wells at concentrations ranging from 1 to 30 mg/mL, followed by incubation for 1 and 3 days. Cell proliferation was determined by adding 20 µL of MTS reagent (BioVision, Milpitas, CA, USA) stock solution per well. The plate was incubated for 2 h at 37 • C in a humidified, 5% CO 2 atmosphere. The plates were then read at 495 nm using a plate spectrophotometer (Multiskan GO, Thermo Fisher Scientific, Waltham, MA, USA). All samples were measured in triplicate. Data on K562 cells viability after the treatment for 24 h and 72h are given at Supporting Information, (Tables S3 and S4).

Statistical Analysis
Statistical processing of results was performed using Statistica 6.0. All data from the three independent experiments were used for measuring the means ± standard deviation (mean ± SD), which were compared using Student's t-test or a nonparametric Wilcoxon Mann-Whitney U test.

Conclusions
In summary, we have shown for the first time the possibility of generating azomethine ylides from 11H-benzo [4,5]imidazo [1,2-a]indol-11-one and α−amino acids, and we also studied their 1,3-dipolar cycloaddition with cyclopropenes and maleimides. As a result of these reactions, novel spiro[benzo [4,5]imidazo[1,2-a]indole-11,2'-pyrrolidine] frameworks were obtained in moderate to good yields, albeit with low stereoselectivity. With the use of a DFT computational study, the observed poor diastereoselectivity was found to result from the formation of two isomeric azomethine ylides (S-and W-shaped dipoles), which subsequently react with cyclic dipolarophiles in a fully diastereoselective fashion. The results of antiproliferative activity study showed that the spiroadducts with a Nisopropylcarbamoyl group at the cyclopropane moiety were more active, while replacement of this group with a phenyl one led to significant decrease in the activity. It was found that, among the tested compounds, spiroadducts 4f and 6i demonstrated a significant activity, with IC 50 5 ± 1 and 12 ± 2 µg/mL, respectively. Despite its low stereoselectivity, the availability of numerous olefins and amino acids makes this method useful for accessing new hybrid spiro-heterocyclic systems containing simultaneously benzimidazole, indole, and pyrrolidine moieties. Current efforts are focused on increasing the stereoselectivity of cycloaddition reactions involving these new azomethine ylides and other dipolarophiles, as well as studying the biological properties of the products.