Pyrrolizines: Promising scaffolds for anticancer drugs

Bioorganic & Medicinal Chemistry 22 (2014) 46–53

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Bioorganic & Medicinal Chemistry journal homepage: www.elsevier.com/locate/bmc

Review

Pyrrolizines: Promising scaffolds for anticancer drugs Amany Belal a,b, Bahaa El-Dien M. El-Gendy c,⇑ a b c

Medicinal Chemistry Department, Faculty of Pharmacy, Beni-Suef University, Beni-Suef 62514, Egypt Pharmaceutical Chemistry Department, Faculty of Pharmacy, Taif University, Taif 21974, Saudi Arabia Chemistry Department, Faculty of Science, Benha University, Benha 13518, Egypt

a r t i c l e

i n f o

Article history: Received 15 September 2013 Revised 12 November 2013 Accepted 20 November 2013 Available online 1 December 2013 Keywords: Pyrrolizines Pyrimido pyrrolizines Thieno pyrrolizines Pyrrolo pyrrolizine Indolo pyrrolizine Perhydropyrrolizines Anticancer drugs SAR

a b s t r a c t Pyrrolizine derivatives constitute a class of heterocyclic compounds which can serve as promising scaffolds for anticancer drugs. The unique antitumor properties of mitomycin C inspired chemists to develop different pyrrolizine systems and assess their potential antitumor activities against a wide variety of cancer types. Here we review the different classes of pyrrolizines that possess anticancer potency, with an emphasis on their structure activity relationships, in an effort to pave the way for further development in this promising area of research. Ó 2013 Elsevier Ltd. All rights reserved.

Contents 1. 2.

3.

Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Pyrrolizines with anticancer activities . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.1. Substituted pyrrolizines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.2. Pyrimido pyrrolizines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.3. Thieno pyrrolizines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.4. Pyrrolo pyrrolizines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.5. Indolo pyrrolizines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 2.6. Perhydropyrrolizines . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . Conclusion . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . References and notes . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .

1. Introduction Pyrrolizines are hetrocyclic systems consisting of two fused five memebered rings with one nitrogen atom at the ring junction; the parent compound of this class is 3H-pyrrolizine 1 (Fig. 1).1 Although pyrrolizine does not appear to occur naturally, many of its derivatives have been isolated from plants2 and animals.3 Pyrrolizidine constitutes the main skeleton of over 660 alkaloids identified in 6000 plants worldwide.4 These alkaloids are ⇑ Corresponding author. Tel.: +20 15 188 75135; fax: +20 13 332 22578. E-mail address: [email protected] (B.E.-D.M. El-Gendy). 0968-0896/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.bmc.2013.11.040

7 8 6

46 47 47 50 50 51 51 52 52 52

1

N 5 4 3 1

2

Figure 1. Structure and numbering system of 3H-pyrrolizine 1.

biosynthesized by plants as secondary metabolites against herbivores.5 Pyrrolizidine alkaloids are known to cause hepatotoxicity and genotoxicity because they form reactive pyrrolic metabolites that can bind to DNA, forming DNA- and DNA–protein-cross-links.6,7

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O N

NH

OH

O

H 2N O

Cl

OH

O

NH

O H 2N Mitomycin C ( 2)

Clazamycin A ( 3)

Clazamycin B ( 4 )

N H

O

O

OH O

7

CJ-16,264 (5)

O

O

NC N

N OH

N

O N

8a-b 8a: X= O 8b: X= S

N

O

N OH

N

O

O H

N OH

R NH

NH N

O

O NC

NC

N

NH

R NH

X NH2

Cl

N

NC

9

N OH 10

O

Figure 3. Substituted pyrrolizines 7–10.

O O

OH

NC

CJ-16,367 (6)

Figure 2. Antitumor antibiotics 2–6.

NC

Morpholine Formaldehyde

CN

NH 2

rt

NH

O

7

11

However, pyrrolizidine alkaloids are beyond the scope of this review. Pyrrolizines and pyrrolizidine alkaloids are challenging synthetic targets that have attracted the attention of many synthesis groups because of the unique structural features and interesting biological activity.8–10 Pyrrolizine analogues are of interest to medicinal chemists, because of their biological and pharmacological activities. The pyrrolizine ring constitutes a scaffold for many compounds with diverse biological roles as antitumor11–13 and anti-inflammatory agents.14–16 One of the early discovered antitumor drugs is mitomycin C 2 (Fig. 2), which was isolated from Streptomyces caespitosus or Streptomyces lavendulae and has antitumor antibiotic activity. It is used to treat upper esophageal carcinoma, anal cancers, breast cancers, and superficial bladder tumors.17–19 Mitomycin C 2 is believed to act as a good DNA alkylator which can cross-link DNA with high efficiency and specificity.20,21 It was demonstrated that mitomycins and structurally similar drugs cross-link the exocyclic amino groups of deoxyguanosine residues at the sequence 50 -d(CG) in duplex DNA.22 Other interesting antitumor antibiotics are clazamycin A 3 and clazamycin B 4. Both compounds are natural pyrrolizidines isolated from microorganisms, the only known pyrrolizidines that contain a chlorine substituent attached to the pyrrolizidine ring.23 Two other well known antibiotics that have a pyrrolizidinone skeleton are CJ-16,264 5 and CJ-16,367 6 (Fig. 2).24 The structural similarity of these compounds shows clearly the importance of the pyrrolizine scaffold as a key nucleus in the synthesis of antitumor drugs. This review summarizes the efforts of medicinal chemists in the search for new pyrrolizine derivatives of potential antitumor activity with an emphasis on their structure activity relationship (SAR) and general synthetic protocols for the major pyrrolizine classes.

N OH

N

Scheme 1. Synthesis of pyrrolizine 7.

O O

O N

O

O

O

OH

OH Combretastatin A-4 13

12

Figure 4. Phenethylidenepyrrolizinones (12) and combretastatin A-4 (13).

Cl Ph

Cl Ph

Ph

N

N 16

Cl

COOH

18 Licofelone

N

R

19a-f O 19a R = H 19d R = OCOC2H5 19b R = OCHO 19e R = OCO-p-C6H4Cl 19c R = OCOCH3 19f R = OCO-m-C6H4-CH3

Cl

Cl

Ph

Ph

N 20

N O

N N

N 21a-c

COO(CH2)nONO2

21a n = 3 21b n = 4 21c n = 5

Figure 5. Licofelone (18) and licofelone derivatives (16, 19–21).

2. Pyrrolizines with anticancer activities 2.1. Substituted pyrrolizines Novel pyrrolizines 7–10 (Fig. 3) were synthesized and tested against mammary cancer cell line (MCF-7) and were found to have higher antitumor activity against breast cancer than the drug doxorubicin. The p-methoxy benzylidene derivative 9 was found to be the most active compound with an IC50 of 16 nM.25 Compound 7 was easily synthesized by stirring 2-(pyrrolidin-2-ylidene)malononitrile 11, morpholine, and formaldehyde at room temperature for 24 h (Scheme 1). Compounds 8a–b, 9, and 10 were obtained by reacting 7 with isocyanates or isothiocyanates, different acylating agents, and p-substituted benzaldehydes respectively.25

Phenethylidenepyrrolizinone 12 (Fig. 4) has a significant cytotoxicity in human KB cells (IC50 = 70 nM). However, the activity of 12 was diminished by replacing either the m-OH or the p-OMe with other substituents. The structural similarity between 12 and combretastatin A-4 13 is remarkable and may provide an explanation for this loss of activity.26 Licofelone 18 (Fig. 5) was developed as an anti-inflammatory drug and it is currently in clinical trials. This drug showed promising anti-tumor activity by enhancing apoptosis in prostate cancer cells as well as HCA-7 colon cancer cells through the mitochondrial pathway.27,28 Moreover, it was identified as a dual 5-LOX/COX inhibitor of both small intestinal and colon tumorigenesis in APCMin/+ mice.29 This drug has great potential for preventing and treating colon cancer, but it has yet to be explored.

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Cl

O

O

O

CN CN

Br

PhCH2MgBr N

14

O

O

O

Cl

Ph

Toluene

Ph

N H

O EtOH, NaHCO3

N

15

N H

O N

Oxalyl chloride THF 10-15 °C

O O

Cl

Ph

N2H 4.H 2O, KOH Ethylene glycol

18 O Licofelone

O

N H

OH

O N

S

O

NH O

O N H

O

N 25

24

S

O

O

Ph

85 °C to 145 °C

N

N H

N

O

NH

N S 26

NH

O O

O 23

S 22

Cl

O

O

N

16

O

NH

NH

O O N

S

S 27

OH O 17

O

O

HN O

O

NH

Scheme 2. Synthesis of licofelone 18.

Gust and co-workers30 synthesized novel licofelone derivatives (16, 19a–f, 20, and 21a–c) through parallel synthesis and tested their cytotoxicity against MCF-7 and MDA-MB-231 cells. The parent compound, licofelone 18, was synthesized following the synthetic route described in Scheme 2. The synthesis started with condensation of 4-chloro-3,3-dimethylbutyronitrile 14 and benzyl-Grignard followed by ring closure to afford 5-benzyl3,3-dimethyl-3,4-dihydro-2H-pyrrole 15. Cyclization of 15 and 2-bromo-1-(4-chlorophenyl)ethanone in aqueous EtOH/NaHCO3 afforded 6,7-diaryl-2,3-dihydro-1H-pyrrolizine 16. Subjecting 16 to Friedel–Craft acylation conditions gave 17 which underwent Wolff–Kishner reduction to give licofelone 18 (Scheme 2).30 Licofelone 18 and 6-(4-chlorophenyl)-2,2-dimethyl-7-phenyl2,3-dihydro-1H-pyrrolizine 16 showed good activity against MCF-7 cells but poor activity against MDA-MB-231 cells. The most potent compounds were 19a, 19b, 19c, and 19d with growth inhibitions comparable to that of cisplatin. The selectivity for MCF-7 cells over MDA-MB-231 cells depends mainly on the substituent at C-5, probably due to specific interactions with some intracellular targets. The most selective compounds, 19e and 19f, were inactive toward MDA-MB-231 cells, but they inhibited the growth of MCF-7 cancer cells with an IC50 = 3.9 and 3.8 lM, respectively. These results showed that tuning the substituent at C-5 of licofelone analogues can enhance both the efficacy and selectivity as an anti-tumor drug.30 Compound 20 was developed as another licofelone derivative and was tested against mammary (MCF-7 and MDA-MB-231) and colon carcinoma (HT-29) cancer cells. It showed a two-fold increase in activity over both 5-flurouracil (5-FU) and licofelone toward mammary carcinoma cells. Moreover, it was as active as 5-FU and twice as active as licofelone toward colon carcinoma cells.31 Licofelone nitric oxide donor structural analogs 21a–c showed high potency as antiproliferative agents against MCF-7 and MDA-MB231 breast cancer as well as at HT-29 colon cancer cells.32 These analogues were found to exhibit higher cytotoxicity at MDA-MB231 cells than the parent licofelone acetic acid, most likely due to the high levels of nitric oxide which may be affecting the mode of action. Pyrrolizine derivatives 22–28 (Fig. 6) were synthesized as mitomycin analogues and their antileukemic activity was tested in an effort to identify better anti-tumor drugs with lower toxicity.33 These derivatives were tested against L1210 leukemia and only compounds with a 5-(2-thienyl) group were found to have chemotherapeutic activity. Compounds 25 and 26 were the most potent compounds with an efficacy comparable to that of mitomycin at higher doses.33

S

28

N

Figure 6. Pyrrolizine derivatives 22–28.

X

ClOC H N

COOH

X 30

X O N

X= O or S

O

COOH Ac2O

N

O

29 31

32 DMAD

X

X

OH

COOCH3

LiAlH4 N

OH

34 X= O or S

N

COOCH3

33

Scheme 3. Synthesis of diol 34.

Pyrrolizine derivatives 22–28 were all derived from the parent diol 34 which was synthesized according to the protocol depicted in Scheme 3. In this protocol, proline 29 was coupled with acid chloride 30 under Schotten–Baumann conditions to give 31. Treatment of 31 with acetic anhydride gave nonisolable mesoionic oxazolones 32 which was cyclized by heating in dimethyl acetylenedicarboxylate (DMAD) to give the diester 33 via 1,3-dipolar cycloaddition. The diester 33 was reduced using lithium aluminum hydride to afford the diol 34 (Scheme 3).33 A series of bis(hydroxymethyl)- and bis(carbamates) pyrrolo[1,2-a]indole derivatives (Fig. 7) were synthesized as bifunctional DNA interstrand cross-linking agents and were found to have antitumor activity against human tumor xenografts (human breast MX-1 xenograft and prostate adenocarcinoma PC3 xenograft) in vivo. The most interesting analogues, 35a–b and 36a–b, showed complete tumor remission in nude mice bearing human breast carcinoma MX-1 xenograft. Moreover, compound 35b significantly suppressed prostate adenocarcinoma PC3 xenograft.34 Acylated vinylogous carbinolamines such as IPP 37 (Fig. 8) showed antineoplastic activity against different tumors.35 This and similar compounds were not designed as carbamoylating agents but rather to resist esterase-catalyzed hydrolysis. The 3,4dichlorophenyl substituent at position 3 was important to maintain the activity, and removing this group or replacing it with

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OH

O NH

O OH

R2

O

N H

HO

N OH

O R2

N

N N

35a-b R 1 35a R1 = H, R 2= Me 35b R 1 = H, R 2 = Et

36a-b

R1

OligonucleotideAnalogue

O

41

36a R 1 = H, R 2 = 4'-OMePh 36b R1 = H,R2 = 2'-OMePh

Figure 9. Oligonucleotide 41.

Figure 7. Bis(hydroxymethyl)- and bis(carbamates) pyrrolo[1,2-a]indole (35a–b, 36a–b).

O

O

O

O O 2N

OCONHCH(CH3)2 OCONHCH(CH3)2

N

O

OCONHCH(CH3)2

X = F, Y =CF 3SO 3 X = Cl, Y =CF 3SO 3 OCONHCH(CH3)2 X = H, Y = IX = F, Y = IX X = Cl, Y= I Y

N

Cl

N

Cl 37 (IPP)

38

N

O

OH

O

O2N

O O

CH3

O

O

O N 45

44

OCONHCH(CH3)2

N

CH3

HO O

OCONHCH(CH3)2

OCONHCH(CH3)2

N

O

43

42

N OCONHCH(CH3)2

O N

Figure 10. Analogues of dehydroretrosine alkaloid 45. F N

N

I

I F 40

39

N O

O

N

Figure 8. IPP analogues.

DNA

R O N ..

O R

N +

N

N

R 46a-b 46a R = H 46b R = OCF3

R 47a-b 47a R = H 47b R = OCF3

DNA

N O R

O R

R = OCONR 1R 2

Figure 11. Aryl-substituted pyrrolizinones 46a–b and 47a–b.

DNA

N Scheme 4. Proposed pyrrolizines.

mechanism

for

DNA

cross-linking

by

DNA biscarbamate

either bromine or phenylthio group decreased the activity and promoted toxicity when tested in the murine P388 lymphocytic leukemia assay.36 This type of compound lacks solubility in water and is unstable in aqueous mixtures.37 To overcome this problem, Anderson et al.38 synthesized a set of a-halopyridinium pyrrolizines 38 (Fig. 8) as prodrugs and tested them against P388 lymphocytic leukemia in vivo. The pyridinium moiety was included to enhance water solubility and reduce carbamate activity and therefore increase the stability of the drug. a-Fluoropyridinium pyrrolizines 39 and 40 showed remarkable activity against P388 lymphocytic leukemia. The significant antitumor activity of biscarbamate pyrrolizines is mainly attributed to their ability to alkylate nucleic acids in cancer cells and therefore inhibit their growth (Scheme 4). Varying the substituents on the heteroaromatic ring can be used to modulate the reactivity and stability of these compounds.39

Woo et al.40 reported oligonucleotides having the pyrrolizine nucleus to diagnose and treat gene related diseases. These compounds can bond with DNA or RNA to target specific sites. The anticancer activity of 2,3-dihydro-6,7-bis(hydroxymethyl)-lHpyrrolizine-5-[40 -(ethyloxymethylamino)-phenyl]-O-oligonucleotide 41 (Fig. 9) was tested against cancer cells such as A549, SKOV-3, SKMEL-2, XF-498 and HCT-15 and showed good growth inhibition.40 Pyrrolizines 42–44 (Fig. 10) possesses structural similarity with dehydroretrosine alkaloid 45, which is a known bifunctional alkylating agent that exhibits remarkable antitumor activity.41 Pyrrolizine 42 forms interstrand cross-links preferentially at the dinucleotide sequence 50 -CpG instead of 50 -GpC, but it lacks any kind of activation mechanism to amend its alkylating ability.42 1H-2,3-Dihydropyrrolizines 43 and 44 with an electron withdrawing group (–NO2) at the 6 position were designed as a masked bisalkylating agent which can be unmasked only after the reduction of the nitro group. Having the nitro group as a trigger reduces the leaving rates of the appropriate groups at positions 1 and 8 to better control the alkylating mechanism of DNA.43 Aryl-substituted pyrrolizinones 46a–b and 47a–b were designed and synthesized as cytochrome P450 aromatase inhibitors (Fig. 11).44 Compounds 46a, 46b, and 47a were as potent as

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4-hydroxyandrostenedione (4-OHA) with an IC50 of 3 ± 0.6, 1.6 ± 0.4, and 0.65 ± 0.07 lM, respectively. The pyridine nitrogen forms a coordinate bond with the heme iron.44 These compounds can fit in the catalytic site of human cytochrome P450 aromatase in a very similar manner to that of the steroid aromatase inhibitors. In the case of meta substituted pyridine derivatives, compound 46b is more active than compound 46a due to the extra hydrophobic surface provided by the OCF3 group. However, in para substituted pyridine derivatives, compound 47b is less active than 47a because of steric hindrance caused by the OCF3 group.44

H N

CN O

H N

N

O

N N

H N

N

O

O 59

58

Figure 13. Pyrimidopyrrolizines 58 and 59.

CN

2.2. Pyrimido pyrrolizines Pyrimido[4,5-b]pyrrolizines 48–50 (Fig. 12) were found to exhibit antitumor activity in mice with melanoma B-16 (T = 40–50%). Compound 49 was active against carcinoma 755 (T = 34%), and 50 was active against Lewis’ bronchial carcinoma (T = 43%) [T% is an index of tumor growth inhibition].45 Moreover, pyrimido[4,5b]pyrrolizine derivatives 51 and 52 were synthesized and their antitumor activities were studied in rats with Jensen sarcoma (JS) and Walker carcinosarcoma (WCS). They were also studied in mice with Lewis lung carcinoma (LLC), carcinoma-755 (Ca-755), hematoblastoma La (La), MOPS plasmacytoma (MOPS), and sarcoma-37 (S-37). Compounds 51 and 52 were found to inhibit solid tumor growth weakly (30–40%) or moderately (60%), with the 4benzylamino-substituted derivative less active than the 4-alkylmercapto derivative.46 A series of novel pyrimido[4,5-b]pyrrolizines 53–57 (Fig. 12) were synthesized, and their antitumor activities were evaluated. Compounds 53–56 inhibited Jensen’s sarcoma in rats to some extent, but they were more active against adenocarcinoma 755 of the mammary gland in mice. The most potent compounds, 55 and 56, inhibited the growth of these tumors by 73 and 78%, respectively, but they were more toxic than 53 and 54.47 Pyrimido[5,4-a]pyrrolizines derivative 57 was tested against cancer cell line (MCF-7) and was found to have higher antitumor activity against breast cancer than the drug doxorubicin (IC50 = 16 lM and 63 lM, respectively).25 Aly et al. reported synthesis of pyrimido[4,5-b]pyrrolizine 58 and pyrimido[5,4-a]-pyrrolizine 59 derivatives (Fig. 13) and tested their antitumor activity against the MCF-7 cell line. These

OEt OEt

N

5-7 °C, 2h

COOEt

COOEt NaOEt

COOEt

N

N CN

CN 60

CN 61

NC

NH2 62 Me2NCH(OEt)2 DMF, 100 °C

COOEt

COOEt N

RX

N RS

N N

EtOH/NaOH S

N 65

COOEt NH4SH

N

EtOH/H2O NC Me2N rt 63

N H 64

N

Scheme 5. Synthesis of pyrimido[4,5-b]pyrrolizine 65.

compounds showed weak to moderate activity, and pyrimido[4,5-b]pyrrolizine 58 was found to be more active than pyrimido[5,4-a]pyrrolizine 59.48 The synthesis of pyrimido pyrrolizine 65 is given as an illustrative example for the synthesis of this class of compounds (Scheme 5). Acetal 60 was reacted with cyanoacetic ester to give l-cyanomethyl-2-(2-ethoxycarbonyl-2-cyano)methylene pyrrolidine 61. Compound 61 was easily cyclized by the Thrope– Ziegler method into 6-amino-5-cyanopyrrolizidine derivative 62 which was reacted with N,N-dimethylformamide diethyl acetal to give enamine 63. Treatment of enamine 63 with ammonium hydrosulfide gave thioamide 64 which was reacted with alkyl halide in alcoholic sodium hydroxide to give pyrimido[4,5-b]pyrrolizine 65 (Scheme 5).45 2.3. Thieno pyrrolizines

O

O

OEt

OEt

O

N

N N

N

N N

N S

O 49 O

OEt

N

N

O

S 48

O

N N

N

S

OEt

S(CH 2) 3N(CH3) 2

Ph 50

51

OEt CN N

N N

N

N

N

N

N

Ph

HN

Ph 52

Ph

O

53

NH2

N

O

N

N

55

Ph 56

N

O

H

N

S

N

N O

S

N

N

N

N Ph

N

54

Ph

N

N

N

Gong and co-workers49 tested the anti-tumor activities of a series of novel methylthio-, sulfinyl-, and sulfonyl-8H-thieno[2,3b]pyrrolizin-8-oximinos in vitro against Bel-7402 (human liver cancer) and HT-1080 (human fibro sarcoma) cell lines. Many compounds showed good antitumor activities. In particular compound 66 (Fig. 14) (IC50 = 18.2 lM, 8.2 lM) was found to be 2.5- and 3.3-times more active than cisplatin (IC50 = 45.2 lM, 26.7 lM) in Bel-740 and HT-1080 cell lines, respectively. In a further elaboration of this work, Gong and co-workers designed another series of novel alkylthio/sulfinyl-8H-thieno[2,3-b]pyrrolizin-8-oximino

N

N S

O

N O

S N

O

N OH O 57

Figure 12. Pyrimido pyrrolizines (48–57).

O 2N

O 66

67

Figure 14. Methylthio-8H-thieno[2,3-b]pyrrolizin-8-oximino derivatives 66 and 67.

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A. Belal, Bahaa El-Dien M. El-Gendy / Bioorg. Med. Chem. 22 (2014) 46–53

S

O

S

O

N R

N

68

71 S

O

S

O

O

S

O N

72a-b 72a R = H 72b R = Me S

N R O 74a-d MeO 74a R = Et 74c R = i-Pr 74b R = n-Pr 74d R = Bu

70 MR22388 OH

N

R

N

MeO

OH

O

of these compounds was very sensitive to changes of the substituents at the phenyl ring at position 3 of the thieno[2,3-b]pyrrolizin8-ones. For example, changing the ether substituent in 74a–d from ethyl (74a) to propyl (74b), isopropyl (74c) or butyl (74d) reduced the activity at least 10-fold. The presence of a hydroxyl or methoxy group in the 3- or 4-position of the phenyl ring is fundamental for activity. Increasing the steric hindrance on the phenyl ring at the meta position decreases the activity dramatically by distorting the best orientation observed by X-ray for the phenyl ring of the most potent compounds with respect to the tricyclic system. For example, compound 75 is 1000-fold less active than 70.54 This remarkable cytotoxicity was retained when small substituents like bromine or iodine were added to the pyrrole ring. For example, pyrrolo[2,3-b]pyrrolizinone 76 has an IC50 of 17 nM when tested in vitro on KB-cells. On the other hand, having larger substituents or aromatic rings on the pyrrole moiety decreases the activity dramatically.55 The general route for synthesis of thieno pyrrolizine is described in Scheme 6. The amino function of the starting material 77a was reacted with dimethoxytetrahydrofuran in the presence of 4-chloropyridinium hydrochloride in dioxane to give the pyrrole derivative 78a. Heating 78a in excess pyrrolidine under reflux conditions gave the corresponding carboxamide 79a. Cyclization of 79a was achieved using phosphorus oxychloride to give the tricyclic ketone 80a in good yield.54

O N

69

HO

S

MeO

S

73 S

O N

O N

75

MeO

OMe

76 OH

Br

Figure 15. Thieno[2,3-b]pyrrolizin-8-ones 68–76.

derivatives and tested them in vitro against Bel-7402, HT-1080, SGC-7901, and A549 cell lines.50 Compound 67 (Fig. 14) was identified as the most potent compound and was superior to cisplatin in all tested cell lines. Several compounds with antiproliferative activities were discovered by Rault and co-workers (Fig. 15). These compounds were based on the thieno[2,3-b]pyrrolizin-8-one scaffold 68 and were tested in vitro against the L1210 leukemia cell line. Four of the tested compounds, 69–71 and 74a–d, showed inhibitory effects in the nanomolar range (IC50 = 15–430 nM). The most potent compound, MR22388 (70, IC50 = 15 nM) was further tested against a panel of nine tumor cell lines and was shown to have antiproliferative activity in the nanomolar range in all cases. It was also found to alter the assembly reaction of microtubules (IC50 = 2.9 lM) in an inhibitory polymerization tubulin test with deoxypodophyllotoxin as internal reference (IC50 = 2.4 lM).51,52 Recently, MR22388 (70) was shown to be a strong and selective inhibitor of a mutated form of tyrosine kinase called FLT3-ITD. Since FLT3-ITD exists in about 25% of normal-karyotype acute myeloid leukemia (NK-AML) and is linked to a bad prognosis, MR22388 (70) is a very promising lead for treating Acute Myeloid Leukemia (AML).53 A study of the structure–activity relationship of this new class of antitubulin agents revealed that several structural requirements were necessary to maintain the activity at an adequate level. The thiophene ring and phenyl substituent at position 3 are crucial. Removing the phenyl ring as in 72a, or replacing it with a small alkyl group as in 72b, diminished the activity. Moreover, the activity was reduced 10-fold when the aromatic ring was placed in position 2 instead of position 3 (e.g., 73). It was also noted that the activity

2.4. Pyrrolo pyrrolizines Rault and co-workers56 has reported the synthesis and antiproliferative activities of novel pyrrolo[2,3-b]pyrrolizinones 81 (Fig. 16) as analogues of their lead compound, MR22388 (70), with a goal of enhancing the activity. These compounds were less active than their thieno analogues with IC50 values ranging from 8.9 to 64 lM. The biological activity was strongly reduced when the sulfur atom of the thieno series was replaced with the free NH of the pyrrolo series; for example, compound 82, the isomer of lead compound MR22388 (70), was 800 times less potent than the lead compound. The N-substituted pyrrolopyrrolizinones were more active than the un-substituted isomers but less active than the thieno derivatives. Compound 83 reduced the growth of two cell lines (MCF-7 and NCI-H460) to less than 32% and was later tested in full panel of 60 cell lines and showed moderate GI50 values in the selected cell lines.56 The synthesis of pyrrolo pyrolizines can be achieved following the synthetic protocol depicted in Scheme 6 using pyrrole derivative 77b as the starting material.56 2.5. Indolo pyrrolizines Novel pyrrolizino[2,3-b]indol-4(5H)-one ring system (Fig. 17) was synthesized and tested against different tumor cell lines. X

X X COOMe DimethoxyTHF

R1 NH2 77a-c

Reflux, 2 h

R1 NH2

1

Pyrrolidine R N

COOMe 77b: R1

79a-c R

1. POCl 3, 80 °C

N COOMe 77c:

NH2

N N

78a-c

N

O

1

Reflux, 2h

R

S 77a:

R

4-Chloropyridinium HCl

COOMe

2. NaOH (10%), 50 °C

COOMe NH 2

X O R

1

N

80a-c Scheme 6. General synthetic route for thieno-, pyrrolo-, and indolo pyrrolizines 80a–c.

52

A. Belal, B.E.-D.M. El-Gendy / Bioorg. Med. Chem. 22 (2014) 46–53

H N H N

O

O

O

N

O

O

O

O N

N

N N

R

N

R4

R = Het or Ar 81

O

OH

O

82

R O

OH

R3

83

R1 O

OH 1

N R2 89

OH

90

Figure 16. Pyrrolo[2,3-b]pyrrolizinones 81–83.

R1 = HO

Derivatives 84 and 85 showed antitumor activity against the HL60(TB) human leukemia cell line, while derivative 86 was selective against MOLT-4 leukemia, HOP-92, A549/ATCC, NCI-H460 nonsmall cell lung cancer, and CAKI-1 renal cancer cell lines.57 Indolo pyrrolizines can be synthesized following the general synthetic protocol depicted in Scheme 6 starting from indole derivative 77c.57

H HO

Figure 19. Perhydropyrrolizines 89 and 90.

O N

2.6. Perhydropyrrolizines

O H O H

N

OHO

Perhydropyrrolizines 87a–d and 88a–c (Fig. 18) were found to inhibit peptidyl-prolyl cis/trans isomerase (PPIase) Pin1. Racemic derivatives 87a–d inhibited Pin1 with inhibition constants of 15, 44, 16, and 32 lM, respectively. To enhance the potency, optically pure derivatives 88a–c were synthesized, and 88c was the most potent compound, with an inhibition constant of 9 lM. The presence of the phosphate moiety and large aromatic substituents was crucial to achieve good binding, since the phosphate helps in binding to the basic cluster, and the substituted perhydropyrrolizines fill the hydrophobic binding pocket of the enzyme.58 A series of dispiro-pyrrolizidino fused oxindoles 89 and dispiropyrrolizidino fused acenaphthoquinone 90 (Fig. 19) were derived from andrographolide, and their anticancer activities were tested against seven cell lines (CHO, HepG2, HeLa, A-431, MCF-7, MDCK and Caco-2). These compounds showed promising activity compared to andrographolide, and further SAR, toxicity, and mechanistic pathway studies are underway.59 Novel pentacyclic polyketides UCS1025A (91) and UCS1025B (92) (Fig. 20) were isolated from the fungus Acremonium sp. KY4917 as new antitumor antibiotics.60 These two polyketides have a unique furopyrrolizidine ring system and their structures and stereochemistry were elucidated through extensive 2D-NMR

H N

H N

O

O

N

O

N

H 3C N

O

87a-d

R

87a: R= Naphthalen-2-yl, R1 = PhCH287b: R= Ph, R1 = PhCH287c: R= Pentaf luorophenyl, R1= PhCH2 87d: R= Naphthalen-2-yl, R1= Piperonyl

91

O

H 92

Figure 20. UCS1025A (91) and UCS1025B (92).

spectroscopy and X-ray crystallography.61 Both compounds showed antiproliferative activity against human tumor cell lines. UCS1025A (91) is a potential chemotherapeutic agent because of its telomerase inhibitory effect (IC50 of 1.3 lM in a TRAP assay).62 3. Conclusion Since the discovery of mitomycin C, chemists have endeavored to develop anticancer drugs based on the pyrrolizine scaffold. These efforts led to the identification of several synthetic small molecules with promising anticancer activities against a wide variety of cancer types. In this review, we have attempted to shed light on the most promising scaffolds among these anticancer drugs based on primary literature. These anticancer drugs include substituted-, pyrimido-, thieno-, indolo-, pyrrolo- and perhydropyrrolizines. It was hypothesized that many of these drugs act by a cross-linking reaction mechanism, because of their structural analogy with mitomycins. In spite of the interesting work on pyrrolizines as anti-tumor drugs that was described in this review, there is still a great need and opportunity for medicinal chemists to further explore the biological activity of pyrrolizine scaffolds through extensive SAR efforts.

86

OH O P OH O 1 HN R

N

H

References and notes

O 85

Figure 17. Pyrrolizino[2,3-b]indol-4(5H)-one 84–86.

H

OHO

O

N

O

O 84

O

H O H

H R R1

OH O P OH Ph O HN

N 88a-c

88a: R= H, R1 = H 88b: R= F, R1 = H 88c: R= H, R1= MeO-

Figure 18. Perhydropyrrolizines 87a–d and 88a–c.

O

1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17.

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