Int J Pharm Pharm Sci, Vol 7, Issue 1, 416-420Original Article

SYNTHESIS AND IN-VITRO STUDY OF NOVEL (Z)-1-BENZHYDRYL-4-CINNAMYLPIPERAZINE DERIVATIVES AS POTENTIAL ANTICANCER AGENTS

S. SHIVAPRAKASH, K. R. KIRAN, LATHA DIWAKAR, G. CHANDRASEKARA REDDY*

Vittal Mallya Scientific Research Foundation, # 94/3 & 94/5, 23^rd Cross, 29^th Main, BTM II Stage, Bangalore 560076, India.
Email: gcreddy@vmsrf.org

Received: 07 Nov 2014 Revised and Accepted: 05 Dec 2014

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ABSTRACT

Objective: The objective of this study was to synthesize Z- 1-benzhydryl-4-cinnamylpiperazines by novel stereo selective synthetic method and evaluation of their anticancer properties.

Methods: A series of novel (Z)-1-benzhydryl-4-cinnamylpiperazine derivatives (9a-j) were synthesized, starting from benzophenones in six steps. Wittig condensation of appropriate benzyltriphenyl phosphonium halides with various 1-benzhydryl- 4-(2-ethanal) piperazines (3a-j), and column purification over silica gel afforded pure Z- 1-benzhydryl-4-cinnamylpiperazines.

Results: The structures of newly synthesized compounds 9a-j were established by ¹H & ¹³C NMR and mass spectral analysis. The anticancer potential (MTT assay) of synthesized compounds was tested against human cervical cancer (HeLa) and murine microglial (BV-2) cell lines. Results indicated that the most of the Z-derivatives exhibited moderate to good anticancer activity on both the cell lines over their E- antipodes.

Conclusion: Compound 9i (cis- flunarizine) exhibited exceptionally superior activity against both HeLa and BV-2 cell lines with IC₅₀ value of 13.23±3.51 µM and 23.1±4.12 µM respectively. Hence, this compound may be considered to be a potential lead molecule for further development

Keywords: Benzophenones, Cinnamylpiperazine derivatives, Wittig reaction, Cinnarizine, Anticancer activity.

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INTRODUCTION

In spite of considerable progress in recent years, cancer remains one of the most difficult diseases to treat and is responsible for about 13% of deaths all over the world. This incidence is increasing due to the ageing of the population in most countries, especially in the developed countries. Further metastasis that sets in may cause about 90% of cancer deaths. Currently, surgery and radiotherapy are the methods used in the treatment of cancer. Another effective way frequently preferred for the treatment of cancer is the systemic chemotherapy. In general, 5-fluorouracil (5-FU) in combination with other anti-cancer agents is used in the treatment of the aerodigestive tract, breast, head, and neck, especially in colorectal cancers therapies with oxaliplatin and irinotecan [1-3]. Cytotoxic and anti- hormonal drugs are the main chemotherapeutics used to reduce the proliferation of malignant cells. On the other hand, significant side-effects along with growth of tumor-cell population are often encountered during chemotherapy [4-7]. Strategies have been working out in many laboratories to look for newer chemical agents, one such example is piperazine and piperazine related compounds which displayed the wide variety of biological activities [8, 9] including apoptosis inducing effects on some cancer cells[10]. In addition, some piperazine compounds with substituent group at position 3 on the piperazine ring can strongly act as a selective κ-opioid receptor agonists [11, 12].

Many piperazine sulfonamide derivatives exhibit MMP-3 enzyme inhibition and carbonic anhydrase inhibition activities [13, 14]. In many cases, piperazine derivatives reduce growth inhibition in human erythroleukemia K562 cells and myeloid leukemia HL-60 cells [15]. Also, inhibit topoisomarase II activity [16].Sampson J. J. et al. reported that, some piperazine derivatives induce apoptosis in U937 cells [17]. The N-Alkyl, N-sulfonyl and N-benzoyl substitution of benzhydrylpiperazine derivatives showed antimicrobial and anticancer properties[18-20]. In few occasions’ compounds with cinnamoyl group often employed in the design of anticancer drugs [21, 22].Encouraged by these literature data and therapeutic value of the piperazine scaffolds prompted us to synthesize some novel piperazine compounds having 1, 4-disubstitutions. Herein, we synthesized various (Z)-1-benzhydryl-4-cinnamylpiperazine derivatives and studied their in-vitro anticancer activity.

MATERIALS AND METHODS

Chemistry

All chemicals purchased were of LR grade from Sigma-Aldrich, Merck, and Loba-chemie; solvents used were of the commercial grade. Melting points were determined on Acro melting point apparatus (using a calibrated thermometer). Thin-layer chromatography (TLC) was run on silica gel pre-coated on aluminium sheet (silica gel 60 F₂₅₄. Merck). Chromatographic separation of mixtures was performed in open glass columns packed with silica gel (Merck Grade 7734, 70-230 mesh) and eluted with ethyl acetate/hexane solvent mixture. Analytical HPLC was recorded with Shimadzu (CLASS-VP) equipped with LC-10AT VP high-pressure pumps, a SPD-M10A VP photodiode array detector, a CTO-10AS VP oven and a SCL-10A VP controller (RP column: Atlantis-T3, 5.0 µm, 4.6x150 mm; Mobile phase: 1% ammonium acetate and 0.2% acetic acid in acetonitrile- gradient elution; UV detector: 230 nm). The Mass spectra were recorded on GCMS-QP2010S (direct probe) instrument and high-resolution mass spectral (HRMS) data were obtained on the Micromass Q-Tof micro instrument using electrospray ionization (ESI). ¹H and [13]C NMR spectra were recorded on a Bruker spectrometer 400 MHz and 100 MHz respectively using CDCl₃ as solvent and TMS as an internal reference.

General procedure for the synthesis of benzyl triphenyl phosphonium salts (4-8)

A mixture of appropriate benzyl halides (1.0 equiv) and triphenylphosphine (1.0 equiv) in toluene was heated under reflux for 10-12 h. the reaction mixture was cooled slowly to room temperature; the precipitated solid were filtered under suction. Finally, be washed with toluene and air dried to afford corresponding benzyltriphenyl phosphonium halides 4-8 as white to off- white solid.

General procedure for the synthesis of (Z)-1-[Bis-(4-chlorophenyl)-methyl]-4-(cinnamyl) piperazine (9a)

Compound 1a (3 g, 9.3 mmol) in DMF (6 mL) was added into a mixture of chloroacetaldehyde dimethylacetal (1.28 g, 10.3 mmol), anhydrous K₂CO₃ (1.42 g, 10.3 mmol) and KI (0.08 g, 0.46 mmol) at room temperature. The reaction mixture was then heated under stirring at 85-90 °C for 6-7 h. Progress of the reaction was monitored by TLC. After the completion of reaction, the mixture was quenched into ice water, and extracted with hexane at room temperature. Organic layer was separated, dried over anhydrous sodium sulphate and concentrated under vacuum. The pure viscous oily material 2a, thus obtained after column purification was taken in 48% aq. HBr (15 mL) and stirred at room temperature for 1-1.5 h. The reaction mixture was then quenched into ice water, basified with dilute NaOH solution and extracted with dichloromethane. The compound 3a present in dichloromethane (30 mL) was dried over anhydrous sodium sulfate, benzyltriphenyl phosphonium chloride (4) (4.0 g, 10.4 mmol) was added. The mixture was cooled to 5 °C; t-BuOK (2.6 g, 23.3 mmol) was added under N₂ atmosphere with continuous stirring. After completion of reaction, the mixture was quenched into water. Organic layer was separated, dried over anhydrous sodium sulphate and concentrated under vacuum. The crude was then subjected to column purification over SiO₂ using EtOAc / hexane as an eluent to afford 9a as a viscous liquid. Overall Yield: 2.10 g (51%). ¹H NMR: δ 7.16 - 7.54 (m, 13 H, Ar-H), 6.56 (d, J =11.8 Hz, 1 H), 5.75 (dt, J =11.8, 6.6 Hz, 1 H), 4.17 (s, 1 H), 3.27 (dd, J = 6.6, 1.8 Hz, 2 H), 2.47 (bs, 4 H), 2.39 (bs, 4 H). [13]C NMR: δ 140.79, 137.10, 131.84, 129.30, 129.14, 128.91, 128.79, 128.59, 128.16, 126.91, 74.67, 56.10, 53.38, 51.75. HRMS calculated for C₂₆H₂₇Cl₂N₂ [M+H]⁺ 437.1551; found 437.1551.

(Z)-1-[Bis-(4-methylphenyl)-methyl]-4-(cinnamyl) piperazine (9b)

The procedure was similar to the one as described for 9a, but compound 1b (3 g, 10.7 mmol) was taken as starting material and the benzyltriphenyl phosphonium chloride (4) was used during Wittig reaction. The compound 9b obtained as a viscous liquid. Overall Yield: 2.20 g (52%). ¹H NMR: δ 7.02 - 7.42 (m, 13 H, Ar-H), 6.55 (d, J =11.8 Hz, 1 H), 5.77 (dt, J =11.8, 6.6 Hz, 1 H), 4.13 (s, 1 H), 3.27 (dd, J = 6.6, 1.8 Hz, 2 H), 2.45 (bs, 4 H), 2.42 (bs, 4 H), 2.25 (s, 6 H, Ar-CH₃). [13]C NMR: δ 140.09, 136.36, 131.34, 130.22, 129.45, 129.15, 128.90, 128.14, 127.74, 126.86, 75.65, 56.17, 53.52, 51.89, 21.03. HRMS calculated for C₂₈H₃₃N₂ [M+H]⁺ 397.2644; found 397.2641.

(Z)-1-[(4-Bromophenyl) phenyl methyl]-4-(cinnamyl) piperazine (9c)

The procedure was similar to the one as described for 9a, but compound 1c (3 g, 9.1 mmol) was taken as starting material and the benzyltriphenyl phosphonium chloride (4) was used during Wittig reaction. The compound 9c obtained as a viscous liquid. Overall Yield: 2.0 g (49%). ¹H NMR: δ 7.19 - 7.46 (m, 14 H, Ar-H), 6.56 (d, J =11.8 Hz, 1 H), 5.76 (dt, J =11.8, 6.5 Hz, 1 H), 4.18 (s, 1 H), 3.27 (dd, J = 6.5, 1.8 Hz, 2 H), 2.46 (bs, 4 H), 2.42 (bs, 4 H). [13]C NMR: δ 142.06, 141.90, 137.07, 131.57, 131.42, 129.56, 129.27, 128.86, 128.56, 128.11, 127.82, 127.13, 126.85, 120.62, 75.45, 56.08, 53.38, 51.75. HRMS calculated for C₂₆H₂₈BrN₂ [M+H]⁺ 447.1436; found 447.1433.

(Z)-1-(Diphenylmethyl)-4-(4'-methoxycinnamyl) piperazine (9d)

The procedure was similar to the one as described for 9a, but compound 1d (3 g, 11.9 mmol) was taken as starting material and the benzyltriphenyl phosphonium halide (5) was used during Wittig reaction. The compound 9d obtained as a viscous liquid. Overall Yield: 2.20 g (46%). ¹H NMR: δ 7.38 - 7.40 (m, 4 H, Ar-H), 7.22 - 7.25 (m, 4 H, Ar-H), 7.13 - 7.19 (m, 4 H, Ar-H), 6.85 – 6.88 (m, 2 H, Ar-H), 6.48 (d, J =11.6 Hz, 1 H), 5.67 (dt, J =11.6, 3.2 Hz, 1 H), 4.21 (s, 1 H), 3.80 (s, 3 H, Ar-OCH₃), 3.26 (dd, J = 2.4, 0.8 Hz, 2 H), 2.49 (bs, 4 H), 2.43 (bs, 4 H). [13]C NMR: δ 158.45, 142.75, 130.69, 130.12, 129.79, 128.40, 127.90, 127.80, 126.83, 113.52, 76.16, 56.20, 55.21, 53.47, 51.86. HRMS calculated for C₂₇H₃₁N₂O [M+H]⁺ 399.2436; found 399.2437.

(Z)-1-(Diphenylmethyl)-4-(3', 5'-dimethoxycinnamyl) piperazine (9e)

The procedure was similar to the one as described for 9a, but compound 1e (3 g, 11.9 mmol) was taken as starting material and the benzyltriphenyl phosphonium halide (6) was used during Wittig reaction. The compound 9e obtained as a viscous liquid. Overall Yield: 2.28 g (44%). ¹H NMR: δ 7.39 (d, J =8.0 Hz, 4 H, Ar-H), 7.23 (t, J =3.6 Hz, 4 H, Ar-H), 7.14 (t, J =6.8 Hz, 2 H, Ar-H), 6.49 (d, J =11.6 Hz, 1 H), 6.41 (s, 2 H, Ar-H), 6.36 (s, 1 H, Ar-H), 5.76 (dt, J =11.6, 6.8 Hz, 1 H), 4.21 (s, 1 H), 3.77 (s, 6 H, Ar-OCH₃), 3.26 (d, J = 6.4Hz, 2 H), 2.49 (bs, 4 H), 2.43 (bs, 4 H). [13]C NMR: δ 160.46, 142.72, 138.97, 131.44, 129.73, 128.38, 127.87, 126.82, 107.02, 99.04, 76.13, 56.12, 55.25, 55.23, 53.42, 51.85. HRMS calculated for C₂₈H₃₃N₂O₂ [M+H]⁺ 429.2542; found 429.2539.

(Z)-1-(Diphenylmethyl)-4-(3', 4'-dimethoxycinnamyl) piperazine (9f)

The procedure was similar to the one as described for 9a, but compound 1f (3 g, 11.9 mmol) was taken as starting material and the benzyltriphenyl phosphonium halide (7) was used during Wittig reaction. The compound 9f obtained as a white solid, mp 104-06 °C. Overall Yield: 2.38 g (46%). ¹H NMR: δ 7.39 (d, J =7.2 Hz, 4 H, Ar-H), 7.22 - 7.26 (m, 4 H, Ar-H), 7.13 - 7.17 (m, 2 H, Ar-H), 6.79 – 6.87 (m, 3 H), 5.69 (dt, J =11.6, 6.8 Hz, 1 H), 4.21 (s, 1 H), 3.88 (s, 3 H, Ar-OCH₃), 3.86 (s, 3 H, Ar-OCH₃), 3.26 (dd, J = 6.8, 1.6 Hz, 2 H), 2.52 (bs, 4 H), 2.43 (bs, 4 H). [13]C NMR: δ 148.50, 148.03, 142.75, 131.27, 130.11, 128.40, 127.89, 127.85, 126.85, 121.54, 112.37, 110.83, 76.16, 56.18, 55.86, 55.83, 53.48, 51.89. HRMS calculated for C₂₈H₃₃N₂O₂ [M+H]⁺ 429.2542; found 429.2544.

(Z)-1-(Diphenylmethyl)-4-(3', 4'-methylenedioxycinnamyl) piperazine (9g)

The procedure was similar to the one as described for 9a, but compound 1g (3 g, 11.9 mmol) was taken as starting material and the benzyltriphenyl phosphonium halide (8) was used during Wittig reaction. The compound 9g obtained as a viscous liquid. Overall Yield: 2.47 g (40%). ¹H NMR: δ 7.39 (d, J =6.8 Hz, 4 H, Ar-H), 7.24 (t, J =7.2 Hz, 4 H, Ar-H), 7.14 (t, J =7.2 Hz, 2 H, Ar-H), 6.68 - 6.78 (m, 3 H, Ar-H), 6.44 (d, J =12.0 Hz, 1 H), 5.94 (s, 2 H), 5.67 (dt, J =12.0, 6.4 Hz, 1 H), 4.21 (s, 1 H), 3.24 (dd, J = 6.4, 1.6 Hz, 2 H), 2.49 (bs, 4 H), 2.43 (bs, 4 H). [13]C NMR: δ 147.40, 146.38, 142.75, 131.25, 130.87, 128.40, 128.33, 127.90, 126.84, 122.70, 109.13, 108.01, 100.94, 76.17, 56.16, 53.48, 51.87. HRMS calculated for C₂₇H₂₉N₂O₂ [M+H]⁺ 413.2229; found 413.2230.

(Z)-1-(Diphenylmethyl)-4-(cinnamyl) piperazine (9h)

The procedure was similar to the one as described for 9a, but compound 1h (3 g, 11.9 mmol) was taken as starting material and the benzyltriphenyl phosphonium halide (4) was used during Wittig reaction.

The compound 9h obtained as a white solid, mp 90-92 °C. Overall Yield: 2.30 g (52%). ¹H NMR: δ 7.10 - 7.42 (m, 15 H, Ar-H), 6.55 (d, J =12.0 Hz, 1 H), 5.77 (dt, J =12.0, 6.6 Hz, 1 H), 4.22 (s, 1 H), 3.28 (dd, J = 6.6, 1.80 Hz, 2 H), 2.46 (bs, 8 H). [13]C NMR: δ 142.8, 137.1, 131.6, 129.5, 128.9, 128.4, 128.1, 127.9, 126.9, 126.8, 76.2, 56.2, 53.5, 51.9. HRMS calculated for C₂₆H₂₉N₂ [M+H]⁺ 369.2331; found 369.2335.

(Z)-1-[Bis-(4-fluorophenyl)-methyl]-4-(cinnamyl) piperazine (9i)

The procedure was similar to the one as described for 9a, but compound 1i (3 g, 10.4 mmol) was taken as starting material and the benzyltriphenyl phosphonium halide (4) was used during Wittig reaction. The compound 9i obtained as a pale yellow viscous liquid. Overall Yield: 2. 0 g (46%). ¹H NMR: δ 7.20 - 7.35(m, 9 H, Ar-H), 6.87-6.98 (m, 4 H, Ar-H), 6.50 (d, J =12 Hz, 1 H), 5.76 (dt, J =12.0, 6.6 Hz, 1 H), 4.20(s, 1 H), 3.27 (dd, J = 6.6, 1.8 Hz, 2 H), 2.47 (bs, 4 H), 2.40 (bs, 4 H). [13]C NMR: δ 163.1, 160.6, 138.3, 137.1, 132.6, 132.5, 131.5, 129.3, 129.2, 128.9, 128.2, 126.9, 115.5, 115.3, 74.5, 56.1, 53.4, 51.7. HRMS calculated for C₂₆H₂₇F₂N₂ [M+H]⁺ 405.2142; found 405.2145.

(Z)-1-[(4-Chlorophenyl) phenyl methyl]-4-(cinnamyl) piperazine (9j)

The procedure was similar to the one as described for 9a, but compound 1j (3 g, 10.4 mmol) was taken as starting material and the benzyltriphenyl phosphonium halide (4) was used during Wittig reaction. The compound 9j obtained as a viscous liquid. Overall Yield: 1. 9 g (45%). ¹H NMR: δ 7.13 - 7.37 (m, 14 H, Ar-H), 6.56 (d, J =12.0 Hz, 1 H), 5.76 (dt, J = 12.0, 6.6Hz, 1 H), 4.20 (s, 1 H), 3.27 (dd, J =6.6, 1.8 Hz, 2 H), 2.48 (bs, 4 H), 2.43 (bs, 4 H). [13]C NMR: δ 142.1, 141.4, 137.1, 132.5, 131.4, 129.3, 129.2, 128.9, 128.6, 128.5, 128.1, 127.8, 127.1, 126.8, 75.4, 56.1, 53.4, 51.8. HRMS calculated for C₂₆H₂₈ClN₂ [M+H]⁺ 403.194; found 403.1940.

Anticancer assay

Cell lines were maintained in Dulbecco’s Modified Eagle’s Medium (Sigma- Aldrich Inc., USA) supplemented with 10% fetal bovine serum (Gibco BRL., USA) in a CO₂ incubator at 37 ^oC. The cytotoxicity of the compounds was measured by MTT assay. Two human cancer cell lines HeLa (cervical) and BV-2 (murine microglial) were placed in a 96-well plate at the density of 10,000 cells per well. After 24 h, cells were treated with various concentrations of compounds from 100 μM serially diluted up to 3.13 μM using cinnarizine, flunarizine and clocinizine as positive controls. The cells were further incubated for 48 h, 20 μl of MTT (5 mg/ml stock, Sigma- Aldrich Inc., USA) was added to each well and incubated for another three hours. The purple formazan crystals formed were dissolved by adding 100 µl of DMSO to each well and absorbance was read at 570 nm in a spectrophotometer [Spectra Max 340]. The cell death was calculated as follows:

Cell death =100- [(test absorbance/ control absorbance) x100]

The test result is expressed as the concentration of a test compound which inhibits the cell growth 50% (IC₅₀).

RESULTS AND DISCUSSION

Chemistry

The procedure outlined in Scheme 1 illustrates the synthesis of (Z)-1-diphenylmethyl-4-cinnamylpiperazine derivatives 9a-j starting from the corresponding benzhydrylpiperazines 1a-j. This in turn obtained from appropriate benzophenones in three steps as described in our earlier procedure [23]. The process comprises the reduction of benzophenone with NaBH_4, conversion of alcohol obtained into benzhydryl chlorides using conc. HCl in toluene and finally treatment with anhydrous piperazine gave compounds 1a-j, which are key intermediates for many known drugs. Further, reaction with equimolar amounts of chloroacetaldehyde dimethylacetal and anhydrous K₂CO₃ in DMF gave compounds (2a-j) which on treatment with aq. HBr (48%) at room temperature afforded the corresponding aldehydes (3a-j).

Finally, Wittig reaction of these aldehydes with appropriate benzyltriphenyl phosphonium halides 4-8 (Table 1) in presence of t-BuOK in dichloromethane followed by column purification over silica gel using EtOAc/hexane (1:9) mixture as eluent, afforded mainly (Z)-1-benzhydryl-4-cinnamylpiperazines (9a-j). All the newly synthesized compounds were characterized by ¹H & ¹³C NMR and mass spectral analysis.

Scheme 1: Synthesis of (Z)-1-benzhydryl-4-cinnamylpiperazine derivatives (9a-j). Reagents and conditions: (a) Chloroacetaldehyde dimethylacetal, K₂CO_3, DMF, 85-90 °C, 6-7 h; (b) HBr 48% in water, rt, 1-1.5 h; (c) Wittig salts (4-8); t-BuOK, CH₂Cl₂, 5 °C, ~3-4 h or till completion of reaction by TLC

Table 1: Benzyltriphenyl phosphonium halides 4-8

Compound	Hal	X	Y	Z	Yield# (%)
4	Cl	H	H	H	95
5	Br	H	OMe	H	88
6	Br	OMe	H	OMe	92
7	Cl	OMe	OMe	H	78
8	Cl	3, 4-Methylene dioxy	H	72

^#isolated yield.

Biological studies: Anticancer activity

Further, compounds (9a-j) were subjected for their in vitro anticancer activity against human cervical cancer (HeLa) and murine microglial (BV-2) cell lines using MTT [3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide] assay as per standard protocol [24]. The IC₅₀ value of each compound was calculated by the excel curve software and the results are summarized in table 2. Cinnarizine, Flunarizine and Clocinizine drugs which are (E) – geometrical isomers were taken as internal reference standards and camptothecin (CPT) as positive control. Results from the table indicated that all the Z-derivatives displayed moderate to good anticancer activity on both the cell lines. Against HeLa cell line, the compounds 9f and 9g with vicinal dioxy system (meta-, para- position) on cinnamyl ring displayed poor anticancer activity.

However, these compounds showed good anticancer effect on BV-2 cell line. Similarly, compounds 9d and 9e exhibited moderate to good inhibitory effect on both the cell lines. In the case of compounds 9h, 9i and 9j, which are antipodes of cinnarizine, flunarizine and clocinizine drugs showed good inhibition against both cell lines. Further, compounds 9a, 9b and 9c with no substitution on cinnamyl ring exhibited moderate to good anticancer activity on both the cell lines. It is noteworthy to mention that of all compounds, the difluoro derivative (9i) found to exhibit significant activity against both the cell lines with IC₅₀ values were very close to the standard drug camptothecin.

Table 2: IC₅₀ values of compounds 9a-j on HeLa and BV-2 cell lines

Compound	IC50 (µM)^ ± SD*
	HeLa cell line	BV-2 cell line
9a	42.68 ±4.91	53.14 ±2.92
9b	32.25 ±4.54	40.84 ±3.82
9c	40.15 ±3.73	26.3 ±7.56
9d	42.72 ±3.61	33.81 ±1.65
9e	41.17 ±2.69	36.0 ±3.79
9f	>200	29.35 ±6.7
9g	>200	34.03 ±1.89
9h (cis-Cinnarizine)[23]	34.54 ±5.57	41.62 ±0.58
9i (cis-Flunarizine)[23]	13.23 ±3.51	23.1 ±4.12
9j (cis-Clocinizine)[23]	14.97 ±5.40	37.28 ±0.3
Cinnarizine#	>200	64.99 ±3.09
Flunarizine#	>200	45.7 ±0.27
Clocinizine#	>200	56.0 ±3.14
CPT	9.57±1.17	14.9±1.21

* The values obtained in at least three separate assays done in triplicate ±SD – Standard deviation, ^ The IC₅₀ value defined as the concentration at which 50% survival of cells was observed, ^# Internal reference drugs.

CONCLUSION

In summary, we have synthesized series of novel (Z)-1-benzhydryl-4-cinnamylpiperazine derivatives (9a-j) and evaluated for their in vitro anticancer potential against HeLa and BV-2 cell lines. The preliminary study indicated that newly synthesized Z- derivatives exhibited moderate to good anticancer activity at micro molar level against selected cancer cell lines. Compounds 9h, 9i and 9j, which are antipodes of cinnarizine, flunarizine and clocinizine exhibited good inhibition against both cell lines. Significant finding is that compound 9i (cis-flunarizine) exhibited potent anticancer effect against both HeLa (IC₅₀ = 13.23±3.51 µM) and BV-2 (IC₅₀ = 23.1 ±4.12 µM) cell lines and stands in close proximity with camptothecin. Hence, compound 9i would stand out as a lead molecule for further study.

ACKNOWEDMENT

We express our sincere thanks to Dr. Anil Kush, CEO Vittal Mallya Scientific Research Foundation Bangalore for his keen interest and encouragement.

CONFLICT OF INTERESTS

Declared None

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