molecules Article
Design, Synthesis, Anticancer Evaluation and Docking Studies of Novel Heterocyclic Derivatives Obtained via Reactions Involving Curcumin Rita M. Borik 1, *, Nagwa M. Fawzy 2 , Sherifa M. Abu-Bakr 2 and Magdy S. Aly 3 1 2 3
*
ID
Department of Chemistry, Jazan University, Jazan 45142, Saudi Arabia Department of Natural and Microbial Products, National Research Center, Dokki, Cairo 12622, Egypt;
[email protected] (N.M.F.);
[email protected] (S.M.A.-B.) Genetics & Cell Biology Division, Department of Zoology, Faculty of Science, Beni-Suef University, Beni-Suef 62511, Egypt;
[email protected] Correspondence:
[email protected]; Tel.: +966-596-763-119; Fax: +966-173-245-538
Received: 12 May 2018; Accepted: 2 June 2018; Published: 8 June 2018
Abstract: Curcumin, a widely utilized flavor and coloring agent in food, has been shown to demonstrate powerful antioxidant, antitumor promoting and anti-inflammatory properties in vitro and in vivo. In the present work, synthesis of new heterocyclic derivatives based on Curcumin was studied. Compound 3 was synthesized via the reaction of furochromone carbaldehyde (1) with Curcumin (2) using pipredine as catalyst. Also, novel, 4,9-dimethoxy-5H-furo [3, 2-g] chromen-5-one derivatives 4a–d, 6a–d, 7, 8a–d, 9 and 10 were synthesized by the reactions of furochromone carbaldehyde (1) with different reagents (namely: appropriate amine 3a–d, appropriate hydrazine 5a–d, hydroxylamine hydrochloride, urea/thiourea, malononitrile, malononitrile with hydrazine hydrate). The structure of the synthesized products had been confirmed from their spectroscopic data (IR, 1 H-NMR, 13 C-NMR and mass spectra). In the present investigation, the newly synthesized products were screened using the MTT colorimetric assay for their in vitro inhibition capacity in two human cancer cell lines (hepatocellular carcinoma (HEPG2) and breast cancer (MCF-7) as well as the normal cell line (human normal melanocyte, HFB4) in comparison to the known anticancer drugs: 5-flurouracil and doxorubicin. The anticancer activity results indicated that the synthesized products 4c and 8b showed growth inhibition activity against HEPG2 cell line and synthesized products 4b and 8a showed growth inhibition activity against MCF-7, but with varying intensities in comparison to the known anticancer drugs, 5-flurouracil and doxorubicin. Cyclin dependent kinase 2 (CDK2), a major cell cycle protein, was identified as a potential molecular target of Curcumin. Furthermore, Curcumin induced G1 cell cycle arrest, which is regulated by CDK2 in cancer cells. Therefore, we used molecular modelling to study in silico the possible inhibitory effect of CDK2 by Curcumin derivatives as a possible mechanism of these compounds as anticancer agents. The molecular docking study revealed that compounds 4b, 8a and 8b were the most effective compounds in inhibiting CDk2, and, this result was in agreement with cytotoxicity assay. Keywords: furochromone carbaldehyde; Curcumin; anticancer activity; cytotoxicity; human cancer cell lines; molecular docking
1. Introduction Heterocyclic compounds are considered an exceptionally important class of compounds which play a key role in health care and prescription drug design [1]. Currently, several heterocyclic chemical substances are available commercially as anti-cancer drugs.
Molecules 2018, 23, 1398; doi:10.3390/molecules23061398
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Curcumin is one of the key components of the curcuminoids group. It is the bioactive ingredient Curcumin is one of the key components of the curcuminoids group. It is the bioactive ingredient of turmeric. Curcumin is considered an outstanding source of new medication development [2]. of turmeric. Curcumin is considered an outstanding source of new medication development [2]. Novel Novel studies have shown that curcuminoids have a variety of natural activities, including studies have shown that curcuminoids have a variety of natural activities, including antioxidant [3,4], antioxidant [3,4], anti-inflammatory [5,6], anticancer [7,8] and anti-angiogenesis [9,10] properties, as anti-inflammatory [5,6], anticancer [7,8] and anti-angiogenesis [9,10] properties, as well as medicinal well as medicinal applications, including their use in HIV therapies [11] and against Alzheimer’s applications, including their use in HIV therapies [11] and against Alzheimer’s disease [12]. disease [12]. The non-toxic food origin and wide range of pharmaceutical drug properties of The non-toxic food origin and wide range of pharmaceutical drug properties of curcuminoids ensure curcuminoids ensure they are promising lead molecules for medicinal chemistry. Efforts have taken they are promising lead molecules for medicinal chemistry. Efforts have taken place to synthesize new place to synthesize new curcuminoid derivatives with better biological activities [13–15]. curcuminoid derivatives with better biological activities [13–15]. On account of these facts, we aimed to prepare novel heterocyclic compounds with different On account of these facts, we aimed to prepare novel heterocyclic compounds with different methods from natural products, namely: Curcumin and formyl furochromone. The newly methods from natural products, namely: Curcumin and formyl furochromone. The newly synthesized synthesized compounds were screened for their in vitro growth inhibition characterization. The in compounds were screened for their in vitro growth inhibition characterization. The in vitro vitro antiproliferative activity of each compound in the study has been determined using the MTT antiproliferative activity of each compound in the study has been determined using the MTT colorimetric assay [16,17] in hepatocellular carcinoma cell line HEPG2 and breast carcinoma cell line colorimetric assay [16,17] in hepatocellular carcinoma cell line HEPG2 and breast carcinoma cell MCF-7 as well as the normal cell line (human normal melanocyte, HFB4). line MCF-7 as well as the normal cell line (human normal melanocyte, HFB4). Computational methods such as molecular docking is very useful and reasonably reliable for Computational methods such as molecular docking is very useful and reasonably reliable for prediction of putative binding modes and affinities of ligands for macromolecules. Such methods are prediction of putative binding modes and affinities of ligands for macromolecules. Such methods are gaining popularity because the experimental determination of complex structures is rather difficult gaining popularity because the experimental determination of complex structures is rather difficult and and expensive. Over the years, the speed and accuracy of computational docking methods has expensive. Over the years, the speed and accuracy of computational docking methods has improved, improved, and these methods now play a significant role in structure-based drug design. and these methods now play a significant role in structure-based drug design. 2. Results and Discussion 2. Results and Discussion 2.1. Chemistry Chemistry 2.1. In an an initial initial study, study, aa typical typical Knoevenagele Knoevenagele Doebner Doebner condensation condensation procedure procedure was was performed, performed, In which involved involved heating heating under under reflux reflux for for aa few few hours hours an an equimolar equimolar quantity quantity of of the the furochromone furochromone which carbaldehyde (1), Curcumin (2) and a base such as pipredine (a few drops) in pyridine as a solvent carbaldehyde (1), Curcumin (2) and a base such as pipredine (a few drops) in pyridine as 3 (Scheme 1) was thus obtained and identified by spectral data such its as IR a[18,19]. solventCompound [18,19]. Compound 3 (Scheme 1) was thus obtained and identified by spectral dataas such spectrum, which showed the disappearance of (CHO gp) and the appearance of (OH gp). its IR spectrum, which showed the disappearance of (CHO gp) and the appearance of (OH gp).
Scheme 1. 1. Synthesis Synthesis of of compound compound 3. 3. Scheme
Multicomponent reactions (MCRs) are frequently used in synthetic, medicinal, and Multicomponent reactions (MCRs) are frequently used in synthetic, medicinal, and combinatorial combinatorial chemistry [20,21]. DHP derivatives [22,23] are an important class of heterocycles due chemistry [20,21]. DHP derivatives [22,23] are an important class of heterocycles due to their biological to their biological pharmacological activities. The reaction conditions were optimized after pharmacological activities. The reaction conditions were optimized after attempting reactions of attempting reactions of equimolar amounts of the furochromone carbaldehyde (1) Curcumin (2) and various amines 3a–d, namely: 3-amino-5-methylisoxazole, thiazol-2-amine, p-toluidine and 1-
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equimolar amounts of the furochromone carbaldehyde (1) Curcumin (2) and various amines 3a–d, Molecules 2018, 23, x FOR PEER REVIEW 3 of 17 namely: 3-amino-5-methylisoxazole, thiazol-2-amine, p-toluidine and 1-naphthylamine respectively, in naphthylamine presence of absolute alcohol/TEA. Formation of compounds 4a–d (Schemeof2)compounds was monitored respectively, in presence of absolute alcohol/TEA. Formation 4a–d by TLC using ethyl acetate: petroleum as eluent. (Scheme 2) was monitored by TLC using ethyl acetate: petroleum as eluent. OH O
O
OH
O
O
O
O O
O
O
O
O
O
O
O
HN
O
O
OH
N
O
HN O
O (4a)
O N 5.5 hrs
O
O
O
(4b)
N S
NH2
s 6hr
EtOH/TEA O
O
O
100 C
O
O
O
O
+ HO
(1)
O
OH
N
S
NH2
EtOH/TEA
OH
(2)
O
100 C
NH2 NH2
O
O
O
O O
O O
O
O O
O O
O HN
(4d)
OH
s hr
6
15
OH
s hr
O O
O O
HN O
OH
OH (4c)
Fig 2: synthesis of compounds (4a-d)
Scheme 2. 2. Synthesis Synthesis of Scheme of compound compound4a–d. 4a–d.
Heterocyclic compounds have significant importance due to their therapeutic properties [24]. Heterocyclic compounds have significant importance due to their therapeutic properties [24]. Also, heterocycles containing pyrazole scaffolds [25,26] exhibit some interesting biological activities. Also, heterocycles containing pyrazole scaffolds [25,26] exhibit some interesting biological activities. Herein, the heterocyclic compounds 6a–d containing pyrazole rings (Scheme 3) were obtained via the Herein, the heterocyclic compounds 6a–d containing pyrazole rings (Scheme 3) were obtained via the condensation reaction of hydrazine derivatives 5a–d (namely: hydrazine hydrate, phenyl hydrazine condensation reaction of hydrazine derivatives 5a–d (namely: hydrazine hydrate, phenyl hydrazine 4-nitrophenyl hydrazine and 2,4-nitrophenyl hydrazine) with furochromone carbaldehyde (1) and 4-nitrophenyl and 2,4-nitrophenyl hydrazine) with time. furochromone carbaldehyde (1) and Curcumin (2)hydrazine in the presence of ethanol/TEA for an appropriate Curcumin (2) in the presence of ethanol/TEA for an appropriate time.
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Scheme Scheme3.3.Synthesis Synthesisofofcompounds compounds(6a–d). (6a–d). Scheme 3. Synthesis of compounds (6a–d).
We also report herein a one-pot protocol for the synthesis of compound 7 (Scheme 4) by a threeWe reportherein hereina one-pot a one-pot protocol the synthesis of compound 7 (Scheme 4) by We also report protocol for for the synthesis of compound 7 (Scheme 4) by a threecomponent condensation of furochromone carbaldehyde (1), Curcumin (2) and hydroxylamine acomponent three-component condensation of furochromone carbaldehyde (1), Curcumin (2) and hydroxylamine condensation of furochromone carbaldehyde (1), Curcumin (2) and hydroxylamine hydrochloride in ethanol using TEA as catalyst. hydrochloride hydrochloride in in ethanol ethanol using using TEA TEA as as catalyst. catalyst. HO HO O O O O O O (1) (1)
O O O O
HO HO O O
O O
++
O O O O
(2) (2)
HO HO
++
EtOH/TEA EtOH/TEA H2N OH stirring/ H2N OH stirring/r.t.r.t. 7 hrs 7 hrs HO HO
O O
N N O O O O
O O O O
O O
O O O O (7) (7)
O O Scheme 4. Synthesis of compound (7). Scheme 4. 4. Synthesis Synthesis of of compound compound (7). (7). Scheme
The Biginelli reaction [27–29] is an important MCR [30] because the products exhibit a wide The Biginelli reaction [27–29] is an important MCR [30] because the products exhibit a wide range The of biological activities. An is acid-catalyzed a one-pot Biginelli reaction [27–29] an important Biginelli MCR [30]reaction because involving the products exhibit athree wide range of biological activities. An acid-catalyzed Biginelli reaction involving a one-pot three component condensation of furochromone carbaldehyde (1), Curcumin (2),a and urea/thiourea gave range of biological activities. An acid-catalyzed Biginelli reaction involving one-pot three component component condensation of furochromone carbaldehyde (1), Curcumin (2), and urea/thiourea gave compounds 8a,b (Scheme 5). carbaldehyde (1), Curcumin (2), and urea/thiourea gave compounds 8a,b condensation of furochromone compounds 8a,b (Scheme 5). Nitrogen (Scheme 5). heterocycles [31] are very important class of chemicals for the synthesis of novel drugs. Nitrogen heterocycles [31] are very important class of chemicals for the synthesis of novel drugs. Here, Nitrogen we reportheterocycles a procedure forarethe one-pot combination furochromone (1), [31] very important class of of chemicals for the carbaldehyde synthesis of novel Here, we report a procedure for the one-pot combination of furochromone carbaldehyde (1), Curcumin (2), and malononitrile [32] to synthesis compound 9 (Scheme 6). drugs. Here, we report a procedure for the one-pot combination of furochromone carbaldehyde (1), Curcumin (2), and malononitrile [32] to synthesis compound 9 (Scheme 6). Derivatives of o-aminocarbonitrile are considered important organic6).intermediates because they Curcumin (2), and malononitrile [32][33] to synthesis compound 9 (Scheme Derivatives of o-aminocarbonitrile [33] are considered important organic intermediates because they have many applications in the synthesis of heterocyclic compounds. One MCRs [34] were utilized to Derivatives of o-aminocarbonitrile [33] are considered importantpot organic intermediates because have many applications in the synthesis of heterocyclic compounds. One pot MCRs [34] were utilized to prepare the target 10 in (Scheme 7) by theofreaction of furochromone carbaldehyde (1), Curcumin they have manycompound applications the synthesis heterocyclic compounds. One pot MCRs [34] were prepare the target compound 10 (Scheme 7) by the reaction of furochromone carbaldehyde (1), Curcumin (2), hydrazine hydrate malononitrile. utilized to prepare theand target compound 10 (Scheme 7) by the reaction of furochromone carbaldehyde (1), (2), hydrazine hydrate and malononitrile. Curcumin (2), hydrazine hydrate and malononitrile.
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HO HO HO
O O O O O O
O O O (1) (1) (1)
O O O
O O H2N O+ H + 2N O + H2N O O
(2) (2) (2)
O O O
++ +
(1) (1) (1)
(2) (2) (2)
HO HO HO
O O O
HO HO HO
O O O O O O
O O O (1) (1) (1)
O O O
++ +
O O O O O O
EtOH/TEA EtOH/TEA EtOH/TEA stirring/ r.t. stirring/ 6 hrs r.t. stirring/ 6 hrs r.t. 6 hrs
O O O
O O O
O O O
O O O
(8a,b) (8a,b) (8a,b)
HO HO HO
O O O
O O O
O O O (9) (9) (9)
O O O
HO HO HO
O O O
CN CN CN CN CN CN
O O O
O O O
O O O
CN CN CN
O O O OH OH OH
N N N
Scheme 6. Synthesis of compound (9). Scheme Scheme 6. 6. Synthesis Synthesis of of compound compound (9). (9). Scheme 6. Synthesis of compound (9). O O O
O O O NH O + H NNH22 + NH O + 2N 2 + O + H + H22N
+ + +
(2) (2) (2) HO HO HO
NH NH22 NH2
O O O
HO HO HO
o o o
O O O
O O O
N N N X X X N N N
Scheme 5. 5. Synthesis of compound compound (8a,b). (8a,b). Scheme Synthesis of of Scheme 5. Synthesis compound (8a,b). Scheme 5. Synthesis of compound (8a,b). HO HO HO O O O
O O O
O O O
O O O
O O O
O O O
O O O
X X X
EtOH/TEA EtOH/TEA EtOH/TEA stirring stirring stirring 60OOC / 10 min 60OC / 10 min 60 C / 10 min a, x= O; b, X=S a, x= O; b, X=S a, x= O; b, X=S
+ + +
HO HO HO
O O O
HO HO HO
O O O
O O O
O O O
O O O
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r.t. r.t. r.t. CN CN CN CN CN CN
N N N
EtOH/TEA EtOH/TEA EtOH/TEA stirring/r.t stirring/r.t stirring/r.t 24Hrs 24Hrs 24Hrs
N N N N N N
O O O
(10) (10) (10) O O O
HO HO HO
Scheme 7. Synthesis of compound 10. Scheme 7. Synthesis of compound 10. Scheme Scheme 7. 7. Synthesis Synthesis of of compound compound 10. 10.
All the synthesized compounds were characterized using IR, All the synthesized compounds were characterized using IR, All the synthesized compounds were characterized using IR, spectrometry. spectrometry. All the synthesized compounds were characterized using spectrometry. mass spectrometry.
CN CN CN
O O O
O O O
O O O O O O
O O O
H-NMR, 13C-NMR and mass H-NMR, 13 C-NMR and mass 13C-NMR and mass H-NMR, 1 IR, H-NMR, 13 C-NMR and
1 1 1
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2.2. Anticancer Activity Most of the anti-tumor drugs currently used in chemotherapy are toxic to normal cells and cause toxicity for immune cells, so it is important to minimize doses to the least amount possible as well as try to minimize the side effects of these drugs. Therefore, the identification of new anti-cancer drug with low side effects on the immune system has become an essential goal in many immuno-pharmacology studies [35]. The newly synthesized products were evaluated for their in vitro cytotoxic activity against human hepatocellular carcinoma (HEPG2) and breast carcinoma (MCF-7) cell lines. Doxorubicin (DOX) and 5-fluorouracil (5-FU), which are two of the most effective anticancer agents, were used as reference drugs. Our results showed that some of the newly synthesized products exhibited a moderate to strong growth inhibition activity on the tested cell lines between 0 and 50 µg/mL concentrations in comparison to the reference anticancer drugs. The relationship between surviving fraction and drug concentration was plotted to obtain the survival curve of the two cell lines. The response parameter calculated was the IC50 value, which corresponds to the concentration required for 50% inhibition of cell viability. Table 1 shows the in vitro cytotoxic activity of the synthesized compounds, where some compounds exhibited significant activity compared to the reference drugs. Table 1. IC50 of the newly synthesized products against the two cell lines. Cell Lines
Compound Solvent 1 3 4a 4b 4c 4d 6a (N5007) 6c (N50011) 6d 7 8a 8b 9 10 5-Flurouracil Doxorubicin
MCF-7
HEPG-2
75.78 53 33 33 20 33 37 26 38 40 28 23 53 29 37 13.35
75.78 50 28 26 50 18 22 29 42 42 35 49 23 36 44 14.70
Some of the tested compounds showed remarkable anticancer activity against MCF-7 cancer cells, while compounds 3, 4a, 4c, 4d, 6a, 6c, 6d, 7, 8b, 9 and 10 had no effect on the breast cells. In the same sense, evaluation the anticancer effect of the tested compounds against human liver HEPG2 cancer cell line revealed that although compounds 3, 4a, 4b, 4d, 6a, 6c, 6d, 7, 8a, 9 and 10 had no antiproliferative effect, compounds 3c and 8b showed anticancer activity close to that of the standard drug. Figure 1 shows the cytotoxic activity of the synthesized product 4b and Figure 2 shows the cytotoxic activity of the synthesized product 8a, against breast MCF7 cancer cell line. Figure 3 shows the activity of the compound 4c, and Figure 4 shows the activity of the compound 8b against HEPG2 cell line in comparison to the reference drugs 5-FU and DOX. From the results listed in Table 1, compounds 4b and 8a showed in vitro cytotoxic activity with IC50 values of 20 and 23, respectively, for the MCF-7 cell line. Compounds 4c and 8b showed an in vitro cytotoxic activity with IC50 values of 18 and 23, respectively for the HEPG2 cell line when the cells were subjected to different concentrations of the compounds.
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Additionally, the results revealed that most of the compounds exhibited no activity against the Additionally, results the compounds exhibited no activity against the the Additionally, the results revealedthat thatmost mostofof the compounds exhibited no activity against growth of normal the HFB4 cells.revealed growth of of normal normal HFB4 growth HFB4 cells. cells. 1.2 1.2 y = 7E-06x2 - 0.0022x + 0.9944 2 - 0.0022x + 0.9944 y = 7E-06xR² = 0.9334 R² = 0.9334 y = 9E-05x2 - 0.0067x + 0.9708 2 - 0.0067x + 0.9708 y = 9E-05xR² = 0.7938 y = 7E-05x2 - 0.0058x + 0.9732 R² = 0.7938 2 - 0.0058x + 0.9732 y = 7E-05xR² = 0.7889 R² = 0.7889 Solvent Solvent y = 0.0001x2 - 0.0174x + 0.9818 Dox 2 - 0.0174x + 0.9818 y = 0.0001xR² Dox = 0.9861 MCF7-4b R² = 0.9861 MCF7-4b HFB4-4b HFB4-4b HEPG2-4b HEPG2-4b
Surviving fraction Surviving fraction
1 1 0.8 0.8 0.6 0.6 0.4 0.4 0.2 0.2 0 0 0 0
y = 0.0004x2 - 0.0344x + 0.9751 2 - 0.0344x + 0.9751 y = 0.0004xR² = 0.9946 R² = 0.9946 10 10
20 20
30 30
40 40
Concentration µg/mL Concentration µg/mL
50 50
60 60
Figure 1. The cytotoxic activity of the synthesized product 4b against breast MCF7 cancer cell line. Figure 1. The cytotoxic activity of the synthesized product 4b against breast MCF7 cancer cell line. Figure 1. The cytotoxic activity of the synthesized product 4b against breast MCF7 cancer cell line.
1.2 1.2 1 1
Surviving fraction Surviving fraction
y = 9E-05x2 - 0.0083x + 0.9801 2 - 0.0083x + 0.9801 y = 9E-05xR² = 0.9501 R² = 0.9501 y = 0.0001x2 - 0.0096x + 0.9991 2 - 0.0096x + 0.9991 y = 0.0001xR² = 0.9975 y = 0.0001x2 - 0.01x +R² 0.9796 = 0.9975 2 y = 0.0001x 0.01x + 0.9796 R² =- 0.9591 Solvent R² = 0.9591 Solvent y = 0.0001x2 - 0.0183x + 0.9809 Fluro 2 - 0.0183x + 0.9809 y = 0.0001xR² = 0.9939 Fluro MCF-7-8a R² = 0.9939 MCF-7-8a HFB4-8a HFB4-8a HEPG2-8a HEPG2-8a y = 0.0005x2 - 0.0383x + 0.9715 2 - 0.0383x + 0.9715 y = 0.0005xR² = 0.9924 R² = 0.9924
0.8 0.8 0.6 0.6 0.4 0.4 0.2 0.2 0 0 0 0
10 10
20 20
30 30
40 40
Concentration µg/mL Concentration µg/mL
50 50
60 60
Figure2.2. The The cytotoxic activity 8a8a against breast MCF7 cancer cellcell line.line. Figure activityof ofthe thesynthesized synthesizedproduct product against breast MCF7 cancer Figure 2. The cytotoxic activity of the synthesized product 8a against breast MCF7 cancer cell line.
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1.2 1.2
Surviving Survivingfraction fraction
1 1
y = 7E-05x2 - 0.007x + 0.9863 y = 7E-05x2 - 0.007x + 0.9863 R² = 0.9718 R² = 0.9718 y = 9E-05x2 - 0.009x + 0.976 y = 9E-05x2 - 0.009x + 0.976 R² = 0.9488 R² = 0.9488 y = 6E-05x22 - 0.009x + 0.9969 y = 6E-05x - 0.009x + 0.9969 Solvent R² = 0.9979 Solvent R² = 0.9979 Fluro Fluro HEPG2-4c HEPG2-4c y = 2E-05x22 - 0.0143x + 0.971 HFB4-4c y = 2E-05x - 0.0143x + 0.971 HFB4-4c R² = 0.9886 MCF7-4c R² = 0.9886 MCF7-4c
0.8 0.8 0.6 0.6 0.4 0.4 0.2 0.2 0 0
y = 0.0004x2 - 0.0355x + 0.9714 y = 0.0004x2 - 0.0355x + 0.9714 R² = 0.993 R² = 0.993 0 0
10 10
20 20
30 30
40 40
50 50
60 60
Concentration µg/mL µg/mL Concentration Figure3.3The The cytotoxic cytotoxic activity against liver HEPG2 cancer cellcell line.line. Figure activity of ofthe thesynthesized synthesizedproduct product4c4c against liver HEPG2 cancer Figure 3 The cytotoxic activity of the synthesized product 4c against liver HEPG2 cancer cell line.
1.2 1.2 y = 7E-06x2 - 0.0021x + 0.98 y = 7E-06x2 - 0.0021x + 0.98 R² = 0.7815 R² = 0.7815
Surviving Survivingfraction fraction
1 1 0.8 0.8
R² = 0.7239 R² = 0.7239
0.6 0.6
Solvent Solvent Dox Dox HEPG2-8b HEPG2-8b HFB4-8b HFB4-8b MCF7-8b MCF7-8b
y = 0.0001x2 - 0.0171x + 0.9831 y = 0.0001x2 - 0.0171x + 0.9831 R² = 0.9918 R² = 0.9918
0.4 0.4 0.2 0.2 0 0
y = 7E-05x2 - 0.0062x + 0.9695 y = 7E-05x2 - 0.0062x + 0.9695 R² = 0.8286 R² = 0.8286
y = 7E-05x2 - 0.0057x + 0.9692 y = 7E-05x2 - 0.0057x + 0.9692
y = 0.0004x2 - 0.0327x + 0.9591 y = 0.0004x2 - 0.0327x + 0.9591 R² = 0.9861 R² = 0.9861 0 0
10 10
20 20
30 30
40 40
50 50
60 60
Concentration µg/mL µg/mL Concentration Figure 4. The cytotoxic activity of the synthesized product 8b against liver HEPG2 cancer cell line. Figure 4. The cytotoxic activity of the synthesized product 8b against liver HEPG2 cancer cell line. Figure 4. The cytotoxic activity of the synthesized product 8b against liver HEPG2 cancer cell line.
2.3. 2.3.Molecular MolecularDocking DockingResults Results 2.3. Molecular Docking Results ToToensure the docked dockedcompounds compoundsrepresented represented favorable valid ensurethat thatthe thebinding binding poses poses of the favorable andand valid To ensure that the binding poses of the docked compounds represented favorable and valid potential parametersand andmethods methodswere werevalidated validated redocking potentialbinding bindingmodes, modes, the the docking docking parameters byby redocking the the potential binding modes, the docking parameters and methods were validated by redocking the cocrystalligand ligandininorder orderto to determine determine the ability of Auto Dock vina toto reproduce thethe orientation andand cocrystal the ability of Auto Dock vina reproduce orientation cocrystal ligand in order to determine the ability of Auto Dock vina to reproduce the orientation and position of theligand ligand observed in in crystal structure. The redocking of cocrystal ligands to their position crystal structure.The Theredocking redocking cocrystal ligands to their positionofofthe the ligandobserved observed in the the crystal structure. of of cocrystal ligands to their respective molecular targets exhibited an RMSD value of
