research article design, synthesis, and molecular docking...

Research ArticleDesign Synthesis and Molecular Docking of1-(1-(4-Chlorophenyl)-2-(phenylsulfonyl)ethylidene)-2-phenylhydrazine as Potent NonazoleAnticandidal Agent

Hazem A Ghabbour1 Maha M Qabeel2 Wagdy M Eldehna3

Abdullah Al-Dhfyan4 and Hatem A Abdel-Aziz15

1Department of Pharmaceutical Chemistry College of Pharmacy King Saud University PO Box 2457 Riyadh 11451 Saudi Arabia2Department of Clinical Microbiology and Immunology College of Medicine Mansoura University Mansoura 35516 Egypt3Department of Pharmaceutical Chemistry College of Pharmacy Egyptian Russian University PO Box 11829 Badr City Cairo Egypt4Stem Cell and Tissue Re-Engineering Program Research Center King Faisal Specialist Hospital and Research CenterPO Box 3354 MBC 03 Riyadh 11211 Saudi Arabia5Department of Applied Organic Chemistry National Research Center Dokki Cairo 12622 Egypt

Correspondence should be addressed to Hazem A Ghabbour ghabbourhyahoocomand Hatem A Abdel-Aziz hatem 741yahoocom

Received 18 October 2014 Revised 14 December 2014 Accepted 14 December 2014 Published 31 December 2014

Academic Editor Tanaji Talele

Copyright copy 2014 Hazem A Ghabbour et al This is an open access article distributed under the Creative Commons AttributionLicense which permits unrestricted use distribution and reproduction in any medium provided the original work is properlycited

1-(1-(4-Chlorophenyl)-2-(phenylsulfonyl)ethylidene)-2-phenylhydrazine (13) was designed and synthesized as potential nonazoleanticandidal agent and precisely characterized by IR 1H NMR 13C NMR and ESI-MS The anti-Candida activity of 13 wasevaluated against four Candida species (C albicans C krusei C parapsilosis and C glabrata) Compound 13 displayed goodanticandidal activities (MIC = 039 0195 039 and 156120583molmL resp) in comparison with that of the standard drug fluconazole(MIC = 0195 inactive 156 and 156 120583molmL resp) against C albicans C krusei C parapsilosis and C glabrata respectivelyA molecular modeling of the newly synthesized compound 13 was built in order to investigate its mode of action towards theprospective target cytochrome P450-dependent enzyme lanosterol 14120572-demethylase (PDB-code 1EA1)The docking results showeda similar binding interaction of 13 and fluconazole at the active site of CYT P450 14120572-sterol demethylase Furthermore compound13 showed no cytotoxicity against normal human breast cell line MCF10A

1 Introduction

In the current medical era human fungal infections havestood out as an important clinical threat with serious asso-ciated morbidity and mortality [1] The human immunodefi-ciency virus (HIV) epidemic stem cell transplantation andmodern progression in the fields of solid organ transplan-tation coupled with the advent of novel immunosuppressivedrugs have collectively resulted in dramatic increase in theincidence and diversity of human fungal infections including

those caused by Candida Aspergillus and Cryptococcus spe-cies [2]

In the last few decades Candida has emerged as one ofthe most prevalent fungal pathogens causing both mucosalcandidiasis and invasive candidemia although it is a part ofthe microbiota of gastrointestinal and urogenital tracts skinand mouth in healthy individuals [3] Many endogenous andexogenous predisposing factors such as immunological disor-ders immoderate administration of antimicrobial agents andprolonged use of invasive catheters may result in impaired

Hindawi Publishing CorporationJournal of ChemistryVolume 2014 Article ID 154357 8 pageshttpdxdoiorg1011552014154357

2 Journal of Chemistry

ON

N

Cl

NNH

SOO

SN

N

R

N

NN

OH

N N

R

NN

N

O

R

N

NN

OHNN

N

R

SN

N

R

S

NNH

N

N

SCl

R

R

The title compound

R1

R1

R1

R1 R1

Econazole 1

Miconazole 2

Isoconazole 3

Sulconazole 4 Butoconazole 5

Voriconazole 8

Oxiconazole 6

Fluconazole 7 Zinoconazole 9 13

R=R1=24-di-Cl

R=24-di-Cl R1=4-Cl

R=24-di-Cl R1=26-di-Cl

R=24-di-F R=24-di-F R1=F R=26-di-Cl

R=4-Cl R1=26-di-ClR=4-Cl R1=24-di-Cl R=R1=24-di-Cl

Figure 1 Structures of antifungal drugs 1ndash9 and the title compound 13

host defence against Candida spp and subsequently Can-dida become pathogenic and disseminate into the blood-stream to infect different organs [4]

According to the mode of action there are main sixclasses of antifungal agents fungal ergosterol synthesis inhi-bitors (azoles ketoconazole fluconazole itraconazole vori-conazole and posaconazole) ergosterol disruptors (polyenesantibiotics amphotericin B and nystatin) squalene epox-idase inhibitors (terbinafine) glucan synthesis inhibitors(echinocandins caspofungin and micafungin) nucleic acidsynthesis inhibitors (5-fluorocytosine) and chitin synthesisinhibitors (Nikkomycin and Polyoxins) [5ndash10]

The azole antifungals are the mainstay of therapy usuallyused to treat different Candida infections such as oropha-ryngeal and esophageal candidiasis vulvovaginal candidia-sis candidemia and disseminated candidiasis [11] Azolesexert antifungal activity through inhibition of the fun-gal cytochrome P450-dependent 14120572-lanosterol demethylasethrough the binding of the N3-atom of the azole moiety tothe sixth coordination of heme iron atom of the porphyrinin the substrate binding site of the enzyme [12] Due to thefungistatic action of azoles on Candida spp cells repetitivelyexposed to these antifungals adapt to the drug pressure andeventually become azole resistant [13]

On the other hand azole antifungal agents showed fatalhepatotoxicity [14ndash16] This side effect may be attributableto the imperfect specificity toward fungal enzyme whereCYP51 is a member of the cytochrome P450 superfamilywhich exists not only in fungi but in mammals [17 18] Thelimited success of azoles due to severe resistance and fatalhepatotoxicity has resulted in an urgent need to give muchattention to update and modify drug leads from the point ofview of medicinal chemistry and drug design to fulfill safeand more potent antifungals

Concerning the SAR of azoles antifungals the azole ringhas been evidenced to be one of the most important pharma-cophores for the activity also it is a key toxicophore for thehepatotoxicity of azole antifungal drugs due to the coordina-tion binding of its nitrogen atom to the iron atom of heme[19 20] All of these findings inspired us to design and preparenovel nonazole lead compound with different structuralpharmacophores hoping to obtain potent antifungal agentand separate the antifungal activity from toxicity

Figure 1 summarized the main structural features of aseries of azoles antifungals such as imidazoles 1ndash6 and triazoledrugs 7 and 8 in addition to zinoconazole 9 (Figure 1) In thepresent study the well-known azole antifungals [21 22] werechosen as lead compounds for development of new nonazolescaffold Firstly the heteroaryl rings were replaced with aphenyl ring Since the anticandidal activity of hydrazoneswas extensively reported [23ndash25] we decided to retain thephenylhydrazine moiety of zinoconazole 9

Finally in the present work utilization of sulfone groupwas performed depending on the reported antimicrobialactivity of sulfones [26ndash28] and due to the incorporation ofsulfur in some azole antifungals such as sulconazole 4 andbutoconazole 5 in addition to the incorporation of oxygen inether functions of econazole 1 miconazole 2 isoconazole 3and hydroxyl group of fluconazole 7 and voriconazole 8

In view of the facts mentioned above and as part ofour ongoing effort to prepare biologically active agents [29ndash36] herein we present the design synthesis and preliminarybiological evaluation of 1-(1-(4-chlorophenyl)-2-(phenylsul-fonyl)ethylidene)-2-phenylhydrazine (13) as potent antifun-gal agent based on a novel nonazole scaffold The mode ofaction of compound 13 to the potential receptor cytochromeP450 14120572-sterol demethylase was investigated by moleculardocking

Journal of Chemistry 3

2 Experimental

21 Chemistry

211 General Melting points were determined on a Gal-lenkamp melting point apparatus and are uncorrectedInfrared (IR) spectra were recorded as KBr disks using thePerkin Elmer FT-IR Spectrum BX apparatus NMR spectrawere scanned in DMSO-119889

6on a Brucker NMR spectrometer

operating at 500MHz for 1H and 125MHz for 13C Chemicalshifts are expressed in 120575-values (ppm) relative to TMS as aninternal standard Coupling constants (119869) are expressed inHzD2O was added to confirm the exchangeable protons Mass

spectra weremeasured on anAgilent Triple Quadrupole 6410QQQ LCMS equipped with an ESI (electrospray ionization)source

212 Synthesis of 1-(4-Chlorophenyl)-2-(phenylsulfonyl)ethan-1-one (12) This compound was prepared according to thereported method [37] To a solution of 2-bromo-1-(4-chlorophenyl)ethan-1-one (10) (10mmol) in absolute ethanol(50mL) sodium benzenesulfinate (11) (12mmol) was addedThe mixture was refluxed for 3 h and then left to cool Thereaction mixture was poured into cold water and the solidproduct filtered off washed with water dried and finallyrecrystallized from EtOH to afford compound 12

213 Synthesis of 1-(1-(4-Chlorophenyl)-2-(phenylsulfonyl)eth-ylidene)-2-phenylhydrazine (13) A mixture of the sulfone 12(1mmol) and phenyl hydrazine (1mmol) in ethanol (30mL)and acetic acid (05mL) was added The reaction mixturewas stirred for 6 h The precipitated product was filteredoff washed with ethanol and dried Recrystallization fromethanol to afford the corresponding hydrazone 13 in 72mp 160ndash162∘C IR ] 3341 (NH) cmminus1 1HNMR(DSMO-119889

6) 120575

517 (s 2H CH2) 682ndash685 (m 1H ArH) 712 (d 119869 = 75Hz

2H ArH) 723ndash726 (m 2H ArH) 732 (d 119869 = 8Hz 2HArH) 753ndash756 (m 2H ArH) 759ndash763 (m 1H ArH) 771 (d119869 = 85Hz 2H ArH) 981 (s D

2O exchangeable 1H NH)

13C NMR (DSMO-1198896) 120575 5154 11300 12006 12726 12788

12812 12893 12897 13188 13393 13650 13926 14446 MS(ESI)119898119911 3847 [M]+

214 Synthesis of Compounds 14andashd 15a b and 16a b Thesecompounds were synthesized recently in a previous study[29]

22 Antifungal Activity In this study we used four referencestrains Candida albicans ATCC 90029 Candida parapsilosisATCC 22019 Candida krusei ATCC 14247 and Candidaglabrata ATCC 15126 Antifungal activity of the synthesizedcompounds was determined by the agar well diffusionmethod [38]

A 1 times 106 CFUsmL yeast suspension was prepared foreach isolate in phosphate-buffered saline (PBS) and Mueller-Hinton agar plates were inoculated automatically with aSpiral plater (Autoplate 4000 Spiral Biotech Inc BethesdaMD USA) Subsequently equidistant (1 cm diameter) holeswere made using sterile cork borer in the agar Holes were

filled with 100120583L of the tested compound at concentration(100 120583mol dissolved in 1mL DMSO) 25120583g fluconazole andsolvent were included as positive and negative controlsrespectivelyThe plates were incubated for 24 h at 37∘C After-wards the antimicrobial activity of each newly synthesizedcompound was evaluated by measuring the inhibition zonediameters The experiment was repeated on three separateoccasions and the average zone of inhibition was calculated

23 Minimal Inhibitory Concentration The microdilutionmethod was performed as described by Irobi et al [39] with96-well round-bottom microtiter plates using RPMI 1640medium (Life Technologies New York NY USA) and 2glucose in MOPS buffered to pH 7 Dimethyl sulphoxide(DMSO) was used as solvent for the synthetic compoundsstarting with 100120583mol concentration of all compounds dis-solved in 1mLDMSOand then reduced by successive twofolddilutions of stock solution using a calibrated micropipetteThe final solutions concentrations were 100 50 25 125 625312 156 078 039 and 0195 120583molmL The stock solutionsof the synthetic compounds were dispersed in the assaymedium to obtain appropriate concentrations in wells 1ndash10in each row drug-free medium was dispensed in wells 11and 12 Fluconazole is used as reference drug Sabourauddextrose agarwas used to culture yeast strainsThe inoculumsof microorganisms were prepared from cultures and wereadjusted to 05 McFarland standard the suspensions werefurther diluted in RPMI 1640 medium to yield an inoculumconcentration of approximately 104 CFUmLThe inoculatedplates were incubated for 48 h at 35∘C Triplicate tests wereperformed and the average was taken as final reading TheMIC endpoint was defined as the lowest drug concentrationexhibiting 100 inhibition of growth compared with thecontrol well [40] According to CLSI criteria 100 growthinhibition is defined as clear wells Therefore the readingsat MIC value were similar to the negative control withoutCandida The MIC of fluconazole was determined for eachspecies in parallel as a control and antibiotic-free solvent wasincluded as a negative control

24 Molecular Docking The crystal structure of cytochromeP450 14120572-sterol demethylase (Cyp51) from Mycobacteriumtuberculosis in complexwith fluconazole (PDB 1EA1)was pro-vided from Brookhaven protein data bank (PDB httpwwwrcsborgpdb) and loaded to Molegro Virtual Docker (MVD2013600 [win32]) program fully functional free trial ver-sion with time limiting license [41] The nonbonded oxygenatoms of water present in the crystal structure was removedChemBio3DUltra 10 [42] was used to draw the 3D structuresof different ligands Ligands were further preoptimized usingfree version of Marvinsketch 4113 from Chemaxon Ltd[43] with MM force field and saved in Tripos mol2 fileformat MolDock score functions were used with a 03 A gridresolution The binding sites were defined to any residueswith 10 A distant from the cocrystallized fluconazole in thecomplex crystal structure of the enzyme [44]

25 Apoptotic Evaluation for Compound 13 Apoptoticeffect of compound 13 against noncancerous human breast


SO OO

ClNaSO OBr

O

Cl

NHCl

N N

SO O

H

+

NH2

10 11 12

13

Scheme 1 Synthetic pathway of the title compound 13

epithelial cell line MCF10A by measurement of annexin-Vbinding by flow cytometry was performed according to thereported method with minor modification [45]

3 Results and Discussion

31 Chemistry 1-(4-Chlorophenyl)-2-(phenylsulfonyl)eth-an-1-one (12) was synthesized by the reaction of 2-bromo-1-(4-chlorophenyl)ethan-1-one (10) with sodium benzenesul-finate (11) (Scheme 1) [37] Sulfone 12 was reacted withphenylhydrazine in ethanol at ambient temperature to affordthe title compound 13 (Scheme 1) The IR spectrum of com-pound 13 revealed the appearance of absorption band ofhydrazone NH at 3341 cmminus1 The 1H NMR of hydrazone 13exhibited the signal of the NH group which appeared at 120575981 and the signal of CH

2protons appeared as singlet at 120575

517 whereas its 13C NMR revealed the signal of CH2carbon

at 120575 5154 The MS (ESI) analysis of 13 showed a peak at119898119911 = 3847 [M]+

In a previous study published recently we synthesizedcompounds 14andashd 15a b and 16a b (Scheme 2) investigatedhere [29]

32 Antifungal Activity Compounds 14andashd 15ab and 16ab[29] and the newly synthesized compound 13 were evaluatedin vitro for their antifungal activity by inhibition zone tech-nique using four Candida species (C albicans C krusei Cparapsilosis and C glabrata) The mean values of the inhi-bition zone diameter obtained for compound 13 suggest thatcompound 13 possesses significant antifungal activity againstC albicans C krusei and C parapsilosis (Table 1)

33 Minimum Inhibitory Concentration (MIC) Thereafterminimum inhibitory concentration (MIC) of the compound13 is evaluated in vitro using the twofold serial dilutiontechnique The lowest concentration showing no growthwas chosen as the MIC The results of minimum inhibitoryconcentration were displayed in Table 2

The antifungal activity of 13 was compared with thatof fluconazole a standard antifungal drug Investigations ofthe antifungal activity against C krusei and C parapsilosisindicated that they were the most sensitive species to the

Table 1 Antifungal activity of compounds 13ndash16 inmean inhibitiondiameters (mm)

Compounds C albicans C krusei C parapsilosis C glabrata13 33 39 35 2214a mdash mdash mdash mdash14b mdash mdash mdash mdash14c 14 11 18 1214d mdash mdash mdash mdash15a mdash mdash mdash mdash15b mdash mdash mdash mdash16a mdash mdash mdash mdash16b mdash mdash mdash mdashFluconazole 29 mdash 27 23

Table 2 Minimum inhibitory concentration (MIC) in 120583molmL of13

Compound C albicans C krusei C parapsilosis C glabrata13 039 0195 039 156Fluconazole 0195 mdash 156 156

influence of the compound 13 with MIC values of 0195and 039 120583molmL respectively Also compound 13 almostwas equipotent as fluconazole against C glabrata (MIC =156 120583molmL) On the other hand compound 13 was 2-foldless active thanfluconazole againstC albicanswithMICvalueof 039 120583molmL

34 Structure Activity Relationship for Compound 13 Fromthe SAR point of view the N-phenyl ring and hydrazonefunction of 13 is necessary for its activity The activity ofcompound 13 may be due to the presence of three electronclouds of aryl groups with similar distribution with those offluconazole (two triazole moieties and benzene ring)

35 Molecular Docking Study for Compound 13 Dockingstudy was performed for 13 in order to investigate the pos-sible interactions with cytochrome P450 14120572-sterol demethy-lase from Mycobacterium tuberculosis (Mycobacterium P450DM) The crystallographic structure of the complex betweencytochrome P450 14120572-sterol demethylase from Mycobac-terium tuberculosis (Mycobacterium P450 DM) and flucona-zole (ID 1EA1) was used for the docking study [46]

Figure 2(a) shows the docked reference drug fluconazolein the active site of the enzyme for validation of our dockingprotocol Azole ring is positioned almost perpendicular to theporphyrin plane (cofactor) where the nitrogen of azole ringcoordinated to the heme ironThe distance between nitrogenatom of azole ring in fluconazole and heme ring was 234 AFluconazole revealed a MolDock score of minus16129 In case ofcompound 13 the oxygen of sulfone moiety is perpendicularto the porphyrin plane heme ion with distance 344 A (Figure2(b)) The phenyl ring of 13 occupied the hydrophobicregion above the heme ring and showed good van der Waalsinteractions with heme and amino acids Tyr76 Phe78 Phe83


MeO

NO

SO

O

H

Cl

NN

SO

O

H

H

Cl

NO

SO

O

H

Me

NO

SO

OMe

H

NN

SO

O

H

H

F

O

SO

OMe

Cl

H

Cl

O

SO

OMe

Cl

H

NO

SO

O

H

14b

15a

14a 14d

15b 16a 16b

14c

Scheme 2 Structure of compounds 14andashd 15a b and 16a b [29]

(a) (b)

Figure 2 (a) Binding mode of fluconazole into the binding site cytochrome P450 14120572-sterol demethylase (b) Binding mode of 13 into thebinding site cytochrome P450 14120572-sterol demethylase

and Phe255 Compound 13 showed excellent MolDock scoreminus16587 better than that of fluconazole (minus16129) consistentwith the results of antifungal activity of 13 and fluconazole

It was observed that compound 13 was oriented in thebinding groove of enzyme in such fashion that favors thepossibility of 120587ndash120587 interaction of its three benzene rings withthe hydrophobic amino acid residues of the binding site of theenzyme (Figure 3) The latter binding interactions of 13 arethe same as that of fluconazolewith binding site of the enzyme(Figure 3) In addition the sulfone moiety in 13 occupies thesame position of the hydroxyl group in fluconazole as shownin Figure 3

36 Apoptotic Activity Evaluation for Compound 13 It hasbeen shown that loss of phospholipid asymmetry of theplasmamembrane is an early event of apoptosisThe annexin-V binds to negatively charged phospholipids like phos-phatidylserine During apoptosis the cells react to annexin-Vonce chromatin condenses but before the plasma membraneloses its ability to exclude DAPI Hence by staining cells witha combination of APC annexin-V and DAPI it is possibleto detect nonapoptotic live cells early apoptotic cells and

late apoptotic or necrotic cells Apoptotic effect of compound13 against noncancerous human breast epithelial cell lineMCF10A showed no effect between the treated (10120583M) andthe untreated cells (Figure 4)

4 Conclusion

In conclusion 1-(1-(4-chlorophenyl)-2-(phenylsulfonyl)eth-ylidene)-2-phenylhydrazine (13) was prepared by the reac-tion of 120573-ketosulfone 12 with phenyl hydrazine and itsstructurewas established under the basis of its spectral data Itshowed excellent antifungal activities against Candida fungalspecies The molecular modeling results showed a similarbinding interaction of 13 and fluconazole in the active site ofCYTP-450 The phenylhydrazone moiety plays an importantrole in the antifungal potentiality of 13 In addition com-pound 13 showed no cytotoxicity against noncancerous cellline MCF10A

Conflict of Interests

Theauthors have declared that there is no conflict of interests


N

N

N

Ph78

N

HO N

N

F

F

S

N

NH

Cl

OO

Tyr76 Ph255

Met99Ser252

Ph83

Figure 3 Overlay of compound 13 (blue) and fluconazole (red) showing their 3D and 2D binding interactions in the groove of cytochromeP450 14120572-sterol demethylase

Untreated

Q1 Q2

Q3 Q4

Q1 Q2

Q3 Q4

10120583m

105

104

103

102

101

105104103102101

DA

BI p

acifi

c blu

e-A

DA

BI p

acifi

c blu

e-A

105

104

103

102

101

Annexin-V APC-A105104103102101

Annexin-V APC-A

Figure 4 Measurement of annexin-V binding by flow cytometry Apoptotic effects of compound 13 against noncancerous human breastepithelial cell line MCF10A showed no effect between the treated (10120583M) and the untreated cells

Acknowledgment

The authors would like to extend their sincere appreciation tothe Deanship of Scientific Research at King Saud Universityfor its funding of this research through the Research GroupProject no RGP-VPP-321

References

[1] T Segura A M Puga G Burillo et al ldquoMaterials with fungi-bioinspired surface for efficient binding and fungi-sensitiverelease of antifungal agentsrdquo Biomacromolecules vol 15 no 5pp 1860ndash1870 2014

[2] M A Ghannoum and J R Perfect ldquoHistory of antifungalsrdquoin Antifungal Therapy chapter 1 pp 1ndash10 Informa Healthcare2010

[3] S Schelenz ldquoManagement of candidiasis in the intensive careunitrdquo The Journal of Antimicrobial Chemotherapy vol 61supplement 1 pp i31ndashi34 2008

[4] F Chimenti B Bizzarri A Bolasco et al ldquoSynthesis and bio-logical evaluation of novel 24-disubstituted-13-thiazoles asanti-Candida spp agentsrdquoEuropean Journal ofMedicinal Chem-istry vol 46 no 1 pp 378ndash382 2011

[5] D J Sheehan C A Hitchcock and C M Sibley ldquoCurrentand emerging azole antifungal agentsrdquo Clinical MicrobiologyReviews vol 12 no 1 pp 40ndash79 1999


[6] D Kong M-J Lee S Lin and E-S Kim ldquoBiosynthesisand pathway engineering of antifungal polyene macrolides inactinomycetesrdquo Journal of Industrial Microbiology and Biotech-nology vol 40 no 6 pp 529ndash543 2013

[7] M Nowosielski M Hoffmann L S Wyrwicz et al ldquoDetailedmechanism of squalene epoxidase inhibition by terbinafinerdquoJournal of Chemical Information andModeling vol 51 no 2 pp455ndash462 2011

[8] M B Kurtz and J H Rex ldquoGlucan synthase inhibitors as anti-fungal agentsrdquo Advances in Protein Chemistry vol 56 pp 423ndash475 2001

[9] V T Andriole ldquoCurrent and future antifungal therapy newtargets for antifungal therapyrdquo International Journal of Antimi-crobial Agents vol 16 no 3 pp 317ndash321 2000

[10] W M Holden J S Fites L K Reinert and L A Rollins-SmithldquoNikkomycin Z is an effective inhibitor of the chytrid funguslinked to global amphibian declinesrdquo Fungal Biology vol 118no 1 pp 48ndash60 2014

[11] M Zervos and F Meunier ldquoFluconazole (Diflucan) a reviewrdquoInternational Journal of Antimicrobial Agents vol 3 no 3 pp147ndash170 1993

[12] C A Hitchcock K Dickinson S B Brown E G V Evans andD J Adams ldquoInteraction of azole antifungal antibiotics withcytochrome P-450-dependent 14120572-sterol demethylase purifiedfromCandida albicansrdquo Biochemical Journal vol 266 no 2 pp475ndash480 1990

[13] ZAKanafani and J R Perfect ldquoResistance to antifungal agentsmechanisms and clinical impactrdquo Clinical Infectious Diseasesvol 46 no 1 pp 120ndash128 2008

[14] B A Stuck A Blum A E Hagner T Hummel L Klimek andK Hormann ldquoMometasone furoate nasal spray improves olfac-tory performance in seasonal allergic rhinitisrdquo Allergy vol 58no 11 p 1195 2003

[15] A Legras A-M Bergemer-Fouquet and A-P Jonville-BeraldquoFatal hepatitis with leflunomide and itraconazolerdquoThe Ameri-can Journal of Medicine vol 113 no 4 pp 352ndash353 2002

[16] N Somchit A R Norshahida A H Hasiah A Zuraini M RSulaiman andMMNoordin ldquoHepatotoxicity induced by anti-fungal drugs itraconazole and fluconazole in rats a comparativein vivo studyrdquoHuman and Experimental Toxicology vol 23 no11 pp 519ndash525 2004

[17] J T Slama J L Hancock T Rho L Sambucetti and K ABachmann ldquoInfluence of some novel 119873-substituted azoles andpyridines on rat hepatic CYP3A activityrdquo Biochemical Pharma-cology vol 55 no 11 pp 1881ndash1892 1998

[18] H Wulff M J Miller W Hansel S Grissmer M D Cahalanand K G Chandy ldquoDesign of a potent and selective inhibitorof the intermediate-conductance Ca2+-activated K+ channelIKCa1 a potential immunosuppressantrdquo Proceedings of theNational Academy of Sciences of the United States of Americavol 97 no 14 pp 8151ndash8156 2000

[19] M Plempel ldquoExperiences recognitions and questions in azoleantimycoticsrdquo Japanese Journal of Medical Mycology vol 23 no1 pp 17ndash27 1982

[20] R A Fromtling ldquoOverview of medically important antifungalazole derivativesrdquoClinicalMicrobiology Reviews vol 1 no 2 pp187ndash217 1988

[21] R L Dyer G J Ellames B J Hamill P WManley and A M SPope ldquoSynthesis of (E)-1-(5-chlorothien-2-yl)-2-(1H-imidazol-1-yl)ethanone 26-dichlorophenylhydrazone hydrochloride anovel orally active antifungal agentrdquo Journal of MedicinalChemistry vol 26 no 3 pp 442ndash445 1983

[22] J M H Molina J Losa A M Brocal and A Ventosa ldquoIn vitroactivity of cloconazole sulconazole butoconazole isoconazolefenticonazole and five other antifungal agents against clinicalisolates of Candida albicans and Candida spprdquoMycopathologiavol 118 no 1 pp 15ndash21 1992

[23] A Ayati M Falahati H Irannejad and S Emami ldquoSynthesisin vitro antifungal evaluation and in silico study of 3-azolyl-4-chromanone phenylhydrazonesrdquo DARU Journal of Pharmaceu-tical Sciences vol 20 no 1 article 46 2012

[24] M D Altntop A Ozdemir G Turan-Zitouni et al ldquoSynthesisand biological evaluation of some hydrazone derivatives as newanticandidal and anticancer agentsrdquo European Journal of Medi-cinal Chemistry vol 58 pp 299ndash307 2012

[25] D M Neumann A Cammarata G Backes G E Palmer andB S Jursic ldquoSynthesis and antifungal activity of substituted246-pyrimidinetrione carbaldehyde hydrazonesrdquo Bioorganicamp Medicinal Chemistry vol 22 no 2 pp 813ndash826 2014

[26] C Curti M Laget A O Carle A Gellis and P Vanelle ldquoRapidsynthesis of sulfone derivatives as potential anti-infectiousagentsrdquo European Journal of Medicinal Chemistry vol 42 no6 pp 880ndash884 2007

[27] W Xu J He M He et al ldquoSynthesis and antifungal activity ofnovel sulfone derivatives containing 134-oxadiazole moietiesrdquoMolecules vol 16 no 11 pp 9129ndash9141 2011

[28] A Muralikrishna B C Venkatesh V Padmavathi A PadmajaP Kondaiah and N S Krishna ldquoSynthesis antimicrobial andcytotoxic activities of sulfone linked bis heterocyclesrdquo EuropeanJournal of Medicinal Chemistry vol 54 pp 605ndash614 2012

[29] H A Abdel-Aziz H A Ghabbour M A Bhat and H-K FunldquoMicrowave-assisted synthesis and characterization of certainoximes hydrazones and olefins derived from 120573-keto sulfones rdquoJournal of Chemistry vol 2014 Article ID 532467 6 pages 2014

[30] A M Alafeefy H A Abdel-Aziz D Vullo et al ldquoInhibi-tion of carbonic anhydrases from the extremophilic bacteriaSulfurihydrogenibium yellostonense (SspCA) and S azorense(SazCA) with a new series of sulfonamides incorporatingaroylhydrazone- [124] triazolo [34-b][134] thiadiazinyl-or 2-(cyanophenylmethylene)-134-thiadiazol-3(2H)-yl moi-etiesrdquo Bioorganic amp Medicinal Chemistry vol 22 pp 141ndash1472014

[31] H A Abdel-Aziz T Elsaman A Al-Dhfyan M I Attia KA Al-Rashood and A-R M Al-Obaid ldquoSynthesis and anti-cancer potential of certain novel 2-oxo-N1015840-(2-oxoindolin-3-ylidene)-2 H-chromene-3-carbohydrazidesrdquo European Journalof Medicinal Chemistry vol 70 pp 358ndash363 2013

[32] H A Abdel-Aziz P Ahmad A Kadi K A Al-Rashood HA Ghabbour and H-K Fun ldquoUnexpected ring-opening of 3-aroylbenzo[b]furans at room temperature a new route for theconstruction of phenol-substituted pyrazolesrdquo Tetrahedron Let-ters vol 54 no 26 pp 3424ndash3426 2013

[33] AMAlafeefy S Isik N A Al-Jaber et al ldquoCarbonic anhydraseinhibitors Benzenesulfonamides incorporating cyanoacryl-amide moieties strongly inhibit Saccharomyces cerevisiae 120573-car-bonic anhydraserdquo Bioorganic and Medicinal Chemistry Lettersvol 23 no 12 pp 3570ndash3575 2013

[34] H A Abdel-Aziz T Aboul-Fadl A-R M Al-Obaid MGhazzali A Al-Dhfyan and A Contini ldquoDesign synthesisand pharmacophoric model building of novel substituted nico-tinic acid hydrazones with potential antiproliferative activityrdquoArchives of Pharmacal Research vol 35 no 9 pp 1543ndash15522012


[35] H A Abdel-Aziz H S A El-Zahabi and K M DawoodldquoMicrowave-assisted synthesis and in-vitro anti-tumor activityof 1 3 4-triaryl-5-N-arylpyrazole-carboxamidesrdquo EuropeanJournal of Medicinal Chemistry vol 45 pp 2427ndash2432 2010

[36] H A Abdel-Aziz and A A I Mekawey ldquoStereoselective syn-thesis and antimicrobial activity of benzofuran-based (1E)-1-(piperidin-1-yl)-N

2-arylamidrazonesrdquo European Journal of

Medicinal Chemistry vol 44 no 12 pp 4985ndash4997 2009[37] J Xiang M Ipek V Suri et al ldquo120573-Keto sulfones as inhibitors of

11120573-hydroxysteroid dehydrogenase type I and the mechanismof actionrdquo Bioorganic ampMedicinal Chemistry vol 15 no 13 pp4396ndash4405 2007

[38] C Perez M Pauli and P Bazerque ldquoAn antibiotic assay by theagar well diffusion methodrdquoActa Biologiae et Medicinal Experi-mentalis vol 15 pp 113ndash115 1990

[39] O N Irobi M Moo-Young and W A Anderson ldquoAntimicro-bial activity of Annatto (Bixa orellana) extractrdquo PharmaceuticalBiology vol 34 no 2 pp 87ndash90 1996

[40] A Urzua M Caroli L Vasquez L Mendoza M Wilkensand E Tojo ldquoAntimicrobial study of the resinous exudate andof diterpenoids isolated from Eupatorium salvia (Asteraceae)rdquoJournal of Ethnopharmacology vol 62 no 3 pp 251ndash254 1998

[41] Molegro Virtual Docker (MVD 2013600) Molegro bioinfor-matics solutions 2013 (Danish) httpwwwmolegrocom

[42] S M Kerwin ldquoChemBioOffice ultra 2010 suiterdquo Journal of theAmerican Chemical Society vol 132 no 7 pp 2466ndash2467 2010

[43] Marvinsketch ldquoversion 610 Chemaxon company cheminfor-matics technology products servicesrdquo 2013 httpwwwche-maxoncom

[44] E Banfi G Scialino D Zampieri et al ldquoAntifungal and antimy-cobacterial activity of new imidazole and triazole derivatives Acombined experimental and computational approachrdquo Journalof Antimicrobial Chemotherapy vol 58 no 1 pp 76ndash84 2006

[45] A A O Sarhan A Al-Dhfyan M A Al-Mozaini C N Adraand T Aboul-Fadl ldquoCell cycle disruption and apoptotic activityof 3-aminothiazolo[32-a]benzimidazole-2-carbonitrile and itshomologuesrdquo European Journal of Medicinal Chemistry vol 45no 6 pp 2689ndash2694 2010

[46] L M Podust T L Poulos andM RWaterman ldquoCrystal struc-ture of cytochrome P450 14120572-sterol demethylase (CYP51) fromMycobacterium tuberculosis in complex with azole inhibitorsrdquoProceedings of the National Academy of Sciences of the UnitedStates of America vol 98 no 6 pp 3068ndash3073 2001

Submit your manuscripts athttpwwwhindawicom

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Inorganic ChemistryInternational Journal of

Hindawi Publishing Corporation httpwwwhindawicom Volume 2014

International Journal ofPhotoenergy


Carbohydrate Chemistry

International Journal of


Journal of

Chemistry


Advances in

Physical Chemistry

Hindawi Publishing Corporationhttpwwwhindawicom

Analytical Methods in Chemistry

Journal of

Volume 2014

Bioinorganic Chemistry and ApplicationsHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

SpectroscopyInternational Journal of


The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Medicinal ChemistryInternational Journal of


Chromatography Research International


Applied ChemistryJournal of



Theoretical ChemistryJournal of


Journal of

Spectroscopy

Analytical ChemistryInternational Journal of


Journal of


Quantum Chemistry


Organic Chemistry International

ElectrochemistryInternational Journal of



CatalystsJournal of


ON

N

Cl

NNH

SOO

SN

N

R

N

NN

OH

N N

R

NN

N

O

R

N

NN

OHNN

N

R

SN

N

R

S

NNH

N

N

SCl

R

R

The title compound

R1

R1

R1

R1 R1

Econazole 1

Miconazole 2

Isoconazole 3

Sulconazole 4 Butoconazole 5

Voriconazole 8

Oxiconazole 6

Fluconazole 7 Zinoconazole 9 13

R=R1=24-di-Cl

R=24-di-Cl R1=4-Cl

R=24-di-Cl R1=26-di-Cl

R=24-di-F R=24-di-F R1=F R=26-di-Cl

R=4-Cl R1=26-di-ClR=4-Cl R1=24-di-Cl R=R1=24-di-Cl

Figure 1 Structures of antifungal drugs 1ndash9 and the title compound 13

host defence against Candida spp and subsequently Can-dida become pathogenic and disseminate into the blood-stream to infect different organs [4]

According to the mode of action there are main sixclasses of antifungal agents fungal ergosterol synthesis inhi-bitors (azoles ketoconazole fluconazole itraconazole vori-conazole and posaconazole) ergosterol disruptors (polyenesantibiotics amphotericin B and nystatin) squalene epox-idase inhibitors (terbinafine) glucan synthesis inhibitors(echinocandins caspofungin and micafungin) nucleic acidsynthesis inhibitors (5-fluorocytosine) and chitin synthesisinhibitors (Nikkomycin and Polyoxins) [5ndash10]

The azole antifungals are the mainstay of therapy usuallyused to treat different Candida infections such as oropha-ryngeal and esophageal candidiasis vulvovaginal candidia-sis candidemia and disseminated candidiasis [11] Azolesexert antifungal activity through inhibition of the fun-gal cytochrome P450-dependent 14120572-lanosterol demethylasethrough the binding of the N3-atom of the azole moiety tothe sixth coordination of heme iron atom of the porphyrinin the substrate binding site of the enzyme [12] Due to thefungistatic action of azoles on Candida spp cells repetitivelyexposed to these antifungals adapt to the drug pressure andeventually become azole resistant [13]

On the other hand azole antifungal agents showed fatalhepatotoxicity [14ndash16] This side effect may be attributableto the imperfect specificity toward fungal enzyme whereCYP51 is a member of the cytochrome P450 superfamilywhich exists not only in fungi but in mammals [17 18] Thelimited success of azoles due to severe resistance and fatalhepatotoxicity has resulted in an urgent need to give muchattention to update and modify drug leads from the point ofview of medicinal chemistry and drug design to fulfill safeand more potent antifungals

Concerning the SAR of azoles antifungals the azole ringhas been evidenced to be one of the most important pharma-cophores for the activity also it is a key toxicophore for thehepatotoxicity of azole antifungal drugs due to the coordina-tion binding of its nitrogen atom to the iron atom of heme[19 20] All of these findings inspired us to design and preparenovel nonazole lead compound with different structuralpharmacophores hoping to obtain potent antifungal agentand separate the antifungal activity from toxicity

Figure 1 summarized the main structural features of aseries of azoles antifungals such as imidazoles 1ndash6 and triazoledrugs 7 and 8 in addition to zinoconazole 9 (Figure 1) In thepresent study the well-known azole antifungals [21 22] werechosen as lead compounds for development of new nonazolescaffold Firstly the heteroaryl rings were replaced with aphenyl ring Since the anticandidal activity of hydrazoneswas extensively reported [23ndash25] we decided to retain thephenylhydrazine moiety of zinoconazole 9

Finally in the present work utilization of sulfone groupwas performed depending on the reported antimicrobialactivity of sulfones [26ndash28] and due to the incorporation ofsulfur in some azole antifungals such as sulconazole 4 andbutoconazole 5 in addition to the incorporation of oxygen inether functions of econazole 1 miconazole 2 isoconazole 3and hydroxyl group of fluconazole 7 and voriconazole 8

In view of the facts mentioned above and as part ofour ongoing effort to prepare biologically active agents [29ndash36] herein we present the design synthesis and preliminarybiological evaluation of 1-(1-(4-chlorophenyl)-2-(phenylsul-fonyl)ethylidene)-2-phenylhydrazine (13) as potent antifun-gal agent based on a novel nonazole scaffold The mode ofaction of compound 13 to the potential receptor cytochromeP450 14120572-sterol demethylase was investigated by moleculardocking


2 Experimental

21 Chemistry







6) 120575




13C NMR (DSMO-1198896) 120575 5154 11300 12006 12726 12788

12812 12893 12897 13188 13393 13650 13926 14446 MS(ESI)119898119911 3847 [M]+









SO OO

ClNaSO OBr

O

Cl

NHCl

N N

SO O

H

+

NH2

10 11 12

13





















MeO

NO

SO

O

H

Cl

NN

SO

O

H

H

Cl

NO

SO

O

H

Me

NO

SO

OMe

H

NN

SO

O

H

H

F

O

SO

OMe

Cl

H

Cl

O

SO

OMe

Cl

H

NO

SO

O

H

14b

15a

14a 14d

15b 16a 16b

14c


(a) (b)






4 Conclusion





N

N

N

Ph78

N

HO N

N

F

F

S

N

NH

Cl

OO

Tyr76 Ph255

Met99Ser252

Ph83


Untreated

Q1 Q2

Q3 Q4

Q1 Q2

Q3 Q4

10120583m

105

104

103

102

101

105104103102101

DA

BI p

acifi

c blu

e-A

DA

BI p

acifi

c blu

e-A

105

104

103

102

101


Annexin-V APC-A


Acknowledgment


References




























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


2 Experimental

21 Chemistry







6) 120575




13C NMR (DSMO-1198896) 120575 5154 11300 12006 12726 12788

12812 12893 12897 13188 13393 13650 13926 14446 MS(ESI)119898119911 3847 [M]+









SO OO

ClNaSO OBr

O

Cl

NHCl

N N

SO O

H

+

NH2

10 11 12

13





















MeO

NO

SO

O

H

Cl

NN

SO

O

H

H

Cl

NO

SO

O

H

Me

NO

SO

OMe

H

NN

SO

O

H

H

F

O

SO

OMe

Cl

H

Cl

O

SO

OMe

Cl

H

NO

SO

O

H

14b

15a

14a 14d

15b 16a 16b

14c


(a) (b)






4 Conclusion





N

N

N

Ph78

N

HO N

N

F

F

S

N

NH

Cl

OO

Tyr76 Ph255

Met99Ser252

Ph83


Untreated

Q1 Q2

Q3 Q4

Q1 Q2

Q3 Q4

10120583m

105

104

103

102

101

105104103102101

DA

BI p

acifi

c blu

e-A

DA

BI p

acifi

c blu

e-A

105

104

103

102

101


Annexin-V APC-A


Acknowledgment


References




























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


SO OO

ClNaSO OBr

O

Cl

NHCl

N N

SO O

H

+

NH2

10 11 12

13





















MeO

NO

SO

O

H

Cl

NN

SO

O

H

H

Cl

NO

SO

O

H

Me

NO

SO

OMe

H

NN

SO

O

H

H

F

O

SO

OMe

Cl

H

Cl

O

SO

OMe

Cl

H

NO

SO

O

H

14b

15a

14a 14d

15b 16a 16b

14c


(a) (b)






4 Conclusion





N

N

N

Ph78

N

HO N

N

F

F

S

N

NH

Cl

OO

Tyr76 Ph255

Met99Ser252

Ph83


Untreated

Q1 Q2

Q3 Q4

Q1 Q2

Q3 Q4

10120583m

105

104

103

102

101

105104103102101

DA

BI p

acifi

c blu

e-A

DA

BI p

acifi

c blu

e-A

105

104

103

102

101


Annexin-V APC-A


Acknowledgment


References




























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


MeO

NO

SO

O

H

Cl

NN

SO

O

H

H

Cl

NO

SO

O

H

Me

NO

SO

OMe

H

NN

SO

O

H

H

F

O

SO

OMe

Cl

H

Cl

O

SO

OMe

Cl

H

NO

SO

O

H

14b

15a

14a 14d

15b 16a 16b

14c


(a) (b)






4 Conclusion





N

N

N

Ph78

N

HO N

N

F

F

S

N

NH

Cl

OO

Tyr76 Ph255

Met99Ser252

Ph83


Untreated

Q1 Q2

Q3 Q4

Q1 Q2

Q3 Q4

10120583m

105

104

103

102

101

105104103102101

DA

BI p

acifi

c blu

e-A

DA

BI p

acifi

c blu

e-A

105

104

103

102

101


Annexin-V APC-A


Acknowledgment


References




























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of


N

N

N

Ph78

N

HO N

N

F

F

S

N

NH

Cl

OO

Tyr76 Ph255

Met99Ser252

Ph83


Untreated

Q1 Q2

Q3 Q4

Q1 Q2

Q3 Q4

10120583m

105

104

103

102

101

105104103102101

DA

BI p

acifi

c blu

e-A

DA

BI p

acifi

c blu

e-A

105

104

103

102

101


Annexin-V APC-A


Acknowledgment


References




























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of























































Journal of

Chemistry


Advances in

Physical Chemistry



Journal of

Volume 2014














Journal of

Spectroscopy



Journal of


Quantum Chemistry






CatalystsJournal of

research article design, synthesis, and molecular docking...

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