SYNTHESIS , REACTIONS AND ANTITUMOR EVALUATION OF SOME POLYCONDENSED THIAZOLOPYRIMIDINE DERIVATIVES

Some novel thiazolo[3,2-a]pyrimidines, pyrimidino[1',2':3,2]thiazolo[4,5-b] pyridines,pyrimidino[1'',2'':3',2']thiazolo[4',5':2,3]pyrido-[2,3-d]pyrimidines, pyrimidino [1',2':3,2]thiazolo[4,5-d]pyrimidines, pyrimidino[1',2':3,2]thiazolo[4,5-d][1,2,4] triazolo[3,4b]pyrimidines (2-10) were prepared starting with 2-(arylmethylene) trimethylthiazolo [3,2a]pyrimidin-3-one(2). Also, some, S-alkylated thiazolo[4,5-d] pyrimidine derivatives were synthesized via reaction of 4-substited-7,7,9-trimethyl-1,3,4trihydropyrimidino[1',2':3,2]thiazolo[4,5-d]pyrimidin-2-thione (6b) with different reagents. Furthermore, some of the prepared products were selected and tested for activity against HCT116 and MCF-7 (human colon carcinoma and human breast carcinoma).


CONCLUSION
The newly synthesized compounds 2b and 13 were tested for antitumor activity and they exhibited a high significant anticancer activity against both of HCT 116cell line and MCF-7 cell line (human colon carcinoma cell and human breast carcinoma cell).The results are summarized in Figures.1-4.

EXPERIMENTAL
Melting points were recorded on an electrothermal IA 9100 digital melting point apparatus and were uncorrected.IR spectra (V max in cm -1 ) were recorded on a Shimadzu FT-IR 8300 spectrophotometer using KBr pellets technique. 1H-NMR and 13 C-NMR spectra were recorded using Bruker WM-400 spectrophotometer using DMSO-d 6 as the solvent and TMS as the internal reference (chemist shifts in ppm).The mass spectra were run at 70 eV with a finnigan SSQ7000 spectrophotometer (thermo-instrument system incorporation, USA) Elemental analysis were operated using Mario El Mentar apparatus, Organic microanalysis unit.Elemental analysis and the above spectra were measured the at National Research Center.Pharmacology was carried out in the Regional Center for Mycology & Biotechnology, Al-Azhar University.

Cytotoxicity assay:
The cells were propagated in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% heat-inactivated fetal bovine serum, 1% L-glutamine, HEPES buffer and 50ug/ml gentamycin.All cells were maintained at 37°C in a humidified atmosphere with 5% CO 2 and were subcultured two times a week.
Cell toxicity was monitored by determining the effect of the test samples on cell morphology and cell viability.
Cytotoxicity evaluation using viability assay: For cytotoxicity assay, the cells were seeded in 96-well plate at a cell concentration of 1x 10 4 cells per well in 100µl of growth medium.Fresh medium containing different concentrations of the test sample was added after 24 h of seeding.Serial two-fold dilutions of the tested chemical compound were added to confluent cell monolayers dispensed into 96-well, flat-bottomed microtiter plates (Falcon, NJ, USA) using a multichannel pipette.The microtiter plates were incubated at 37°C in a humidified incubator with 5% CO 2 for a period of 48 h.Three wells were used for each concentration of the test sample.Control cells were incubated without test sample and with or without DMSO.The little percentage of DMSO present in the wells (maximal 0.1%) was found not to affect the experiment.After incubation of the cells for 24 h at 37°C, various concentrations of sample (50, 25, 12.5, 6.25, 3.125 & 1.56 \ig) were added, and the incubation was continued for 48 h and viable cells yield was determined by a colorimetric method.
In brief, after the end of the incubation period, media were aspirated and the crystal violet solution (1%) was added to each well for at least 30 minutes.The stain was removed and the plates were rinsed using tap water until all excess stain is removed.Glacial acetic acid (30%) was then added to all wells and mixed thoroughly, and then the absorbance of the plates were measured after gently shaken on Microplate reader (TECAN, Inc.), using a test wavelength of 490 nm.All results were corrected for background absorbance detected in wells without added stain.Treated samples were compared with the cell control in the absence of the tested compounds.All experiments were carried out in triplicate.The cell cytotoxic effect of each tested compound was calculated (Mosmann, 1983;Vijayan et al., 2004).

Comment: Inhibitory activity against breast carcinoma cells was detected under these experimental conditions with IC 50 = 2.4 µg. Table (1)
Characterization data of compounds 2-17.