I Title:
2-(3,SDimethyl- 1 H-pyrazol- 1 -yl)- 1 -(naphth-2-y1)ethanone as potential anticolon cancer
agent.
Field of Invention:
The present invention relates anticancer screening of the compound, 2-(3,5-Dimethyl- 1 Hpyrazol-
1 -yl)- 1 -(naphth-2-y1)ethanone against colon (HCTI 16 and HT29), prostate (DU 145)
and ovarian (SKOV3) cancer cell lines.
Background of the invention:
Pyrazole derivatives have attracted much attention in recent past owing to their various
biological properties in drug discovery. They act as inhibitors against DNA gyrase and
topoisomerase IV at their respective ATP binding sites, GABA receptor antagonists. Such
compounds also found to exhibit important chemotherapeutics potential like anticancer,
antiproliferative, and antineoplastic etc. Some 3, 5-dimethylpyrazole derivatives having were
found to act as potent antibacterial and anticancer agents. It has already been reported in
literature that some phenacyl and napthacyl substituted azoles exhibit a wide range of
biological activities. For example N-Phenacylpyrazoles act as key precursors for the
discovery of regulators for apoptosis and autophagy in A549 lung cancer cells, N-(4-
substituted phenacy1)imidazoles and N-(4-substituted phenacy1)-2-methylimidazole act as
potent neuronal nitric oxide synthase inhibitors used in the treatment of cerebral ischemia and
neurodegenerative diseases. So, these mentioned facts prompted us to evaluate anticancer
potential of 2-(3,5-Dimethyl- 1 H-pyrazol- 1 -yl)- 1 -(naphth-2-y1)ethanone which was
synthesized in our lab.
3327KOLNPl2007 concerns hydrazinocarbonylthieno[2,3-c] pyrazoles, their preparation
method, compositions containing same and use thereof. In particular, the invention concerns
preparation of hydrazinocarbonylthieno[2,3-clpyrazoles, their preparation method,
compositions containing same, and their use as medicine, in particular as anticancer agents.
1065/CHENP/2011 concerns 3,4-diarylpyrazoles as protein kinase inhibitors whereas the
present invention is related to pyrazoles with methyl groups at positions 3 and 5.
a
1971/KOLNPl2007 is related to pyrazole compounds which are structurally different from
the present invention.
27031 KOLNPl2007 concerns pyrazole compounds useful as protein kinase inhibitors
4681/DELNPl2007 concerns fused pyrazoles wherein the compound claimed is structurally
different from the present invention.
None of the prior art indicates above either alone or in combination with one another disclose
what the present invention has disclosed.
Summary of the invention:
The present invention discloses anticancer screening of some pyrazole based compounds,
against colon (HCT116 and HT29), prostate (DU145) and ovarian (SKOV3) cancer cell lines.
The compound 2-(3,5-Dimethyl-1 H-pyrazol- 1 -yl)- 1 -(naphth-2-y1)ethanone was found to be
cytotoxic to all the cancer cell lines, except SKOV3, with more selectivity towards the colon
cancer cell lines HCTl16 and HT29.
Detailed description of the invention:
Anticancer Activity
To demonstrate the versatility of the synthesized compounds (Fig. I), we evaluated their anticancer
potential in several cancer cell lines. The novel agents were first screened against
colon cancer cell line HCT116. Cells were treated with various concentrations (0, 0.5, 1 , 2.5,
5, 10 ,25, and 50 pM) of the compounds for 48 h. Table 1 shows the % cell viability of four
compounds namely, la, lc, l j and l k while the other compounds tested did not show any
effect on cell viability compared to control in MTT assay. These four compounds were then
selected for screening against colon (HCTI 16 and HT29), prostate (DU 145), and ovarian
(SKOV3) cancer cell lines for a longer time period of 72 h treatment, to establish their anticancer
potential in different cancers. Interestingly, none of the compounds was effective on
SKOV3 cells.
la-k
Fig. 1
Furthermore, compounds la, l c and l k did not have any effect on viability of HT29 and
DU145 cells even at the highest tested concentration (50 pM). Only, compound 2-(3,5-
Dimethyl- 1 H-pyrazol-1 -yl)- 1 -(naphth-2-y1)ethanone 1 j was found to be cytotoxic to colon
and prostate cancer cells, with relatively more pronounced efficacy towards colon cancer
cells (Fig. 2) in 72h cell viability assay. The % cell viability of different cancer cell lines in
presence of increasing dose of l j clearly shows it to be more effective in colon cancer cells,
as compared to prostate and ovarian cancer cells. Hence, l j has the potential to be developed
as a therapeutic agent for colon cancer and warrants further characterization of its efficacy
and mechanism of action.
Table 1. The average mean value (* SD) of percent cell viability (48 h treatment) compared
to the cells treated with DMSO.
Colon cancer (HCT116) cells
Compounds
10 pM 25 pM 50 pM
Fig.2. MTT assay showing the cell viability of colon (HT29 and HCTl16), ovarian (SKOV3)
and prostate (DU145) cancer cell lines after treatment with l j for 72 h. Compound l j
effectively reduced the cell viability of both colon cancer cell lines compared to DMSO
treated cells. The graph shows the average values represented as percentage of control DMSO
treated cells. The error bars represents the SD. Statistics using students t-test for unpaired
values. * p < 0.05 and ** p < 0.005 for control cells versus l j treated cells.
Outcome of the Invention:
Compound lj showed growth inhibitory effects in different cancer cell lines, with more
selectivity towards colon cancer cells compared to ovarian and prostate cancer cells.
Therefore, compound l j has the potential to be developed further as a therapeutic for colon
cancer in future.
Highlights of the Invention:
*:* Explored pyrazole based ethanone l j as a new anticancer agents for colon cancer.
Experimental:
Anticancer activity
Cell Culture
SKOV3 cell lines were grown in DMEM medium (10-013-CV, Corning), HCTl16 and HT29
were maintained in McCoy's 5A medium (166600-082, Life techonlogies) and DU145 were
grown in Eagle's Minimum Essential Medium (EMEM). All the media contained 10% fetal
bovine serum (FBS), 100 units/mL penicillin, and 100 mg/ml streptomycin. Cells were grown
in a 37 "C incubator with 5% COz.
Cell Viability assay
The effect of the compounds on the cell viability of different cancer cell lines was determined
by performing colorimetric MTT assay as previously described, in which the MTT (3-(4,5-
dimethylthiazol-2-y1)-2,5-diphenyletetrazolium bromide) is reduced to its insoluble crystal
form formazan. Briefly, different cancer cells (HCTI 16, HT29, DU 145, and SKOV3) were
incubated in 96 well plates at concentration of 30000 cells per mL in a 96 well plate for
overnight. Further, the cells were treated with increasing concentration of the compounds (O-
50 pM) in triplicates for 48h and 72h time points. MTT (M2128, Sigma) reagent (5 mg/mL)
was added to each well 3h prior to the termination time point and plates were placed back in
the incubator at 37 "C. After 3h, the formazan crystals were dissolved in 100 pL DMSO.
Optical density was measured at 570 nm by using a microplate reader (Dynex Technologies,
MRX Revelation; Chantilly, VA, USA). The background reading was measured at 630 nm,
which was subtracted from 570 nm reading during the cell viability calculation.
5

We claim
1. A compound of the formula 2-(3,5-Dimethyl- 1 H-pyrazol- 1 -yl)- 1 -(naphth-2-
yl)ethanone,
wherein the said compound exhibits cytotoxic activity by reducing cell viability of
colon (HCT116 and HT29) and prostate (DU 145) cancer cell lines in different
concentrations.
2. A compound as claimed in claim 1, wherein the said compound reduces cell viability
of colon (HCT116) cancer cells upto 36.45% in 48 hours cell viability assay.
3. A compound as claimed in claim 1, wherein the said compound is used in
concentrations upto 50 pM.
4. A compound as claimed in claim 1, wherein the said compound reduces cell viability
of colon (HCTI 16) cancer cells upto 57% at 10 pM after 48 hours treatment.
5. A compound as claimed in claim 1, wherein the said compound reduces cell viability
of colon (HCT116) cancer cells upto 56% at 25 pM after 48 hours treatment.
6. A compound as claimed in claim 1, wherein the said compound exhibits selectivity
for the colon cancer cell lines.
7. A compound having same basic skeleton as claimed in claim 1.
Dated this January 17, 2014 6 (Satya Pal Arora)
AUTHORIZED AGENT FOR THE APPLICANT
PATENT AGENT REGISTRATION NO. IN/PA-389