Rahul Raghavan

Tetracycline and Glutathione Inhibit Matrix Metalloproteinase Activity: An In Vitro Study Using Culture Supernatants of L929 and Dalton Lymphoma Cell Lines

Tetracycline and glutathione inhibited the protease activities of matrix metalloproteinase-2 and matrix metalloproteinase-9 expressed by mouse fibrosarcoma cells (L929) and Dalton lymphoma cells, respectively. The inhibitory activity of the tetracycline may be due to its ability to chelate metal ions such as calcium and zinc. Gelatin-zymography technique was used to demonstrate the inhibitory activity of both tetracycline and glutathione. The intensity of the bands corresponding to metalloproteinase activity in zymography gel was reduced in the presence of 50–100 μg/mL of tetracycline. The presence of 10–100 μg/mL of tetracycline in the medium increased the adherence of L929 cancer cells. These results clearly indicate the antimetastatic property of tetracycline. Reduced glutathione, a compound which is produced endogenously by the cells to maintain the redox status, was shown to inhibit the matrix metalloproteinase activity (in vitro). Therefore, it is assumed that decreased glutathione levels in synovial fluids or plasma might increase the activity of MMP. Reduced glutathione at 100 μg/mL inhibited the metalloproteinase activity in gelatin-zymographic gel. As both tetracycline and glutathione exhibited an inhibitory effect on matrix metalloproteinase activity, it was of great interest to check their clinical effects on various MMP associated pathological conditions such as cancer metastasis and arthritis. Here we report that tetracycline and reduced glutathione inhibited the activity of MMP2 completely and activity of MMP9 partly.

Cytotoxic Effect of (Z)-Ethylidene-4,6-Dimethoxycoumaran-3-One Isolated from Pogostemon quadrifolius (Benth.) on PC-3 and DU-145 Prostate Cancer Cells

The study was conducted to identify the antiproliferative property and the mode of action of (Z)-ethylidene-4,6-dimethoxycoumaran-3-one (EDC), a novel compound isolated from the plant leaves of Pogostemon quadrifolius (Benth.), against prostate cancer cell lines PC-3 and DU-145. EDC inhibited the proliferation of PC-3 and DU-145 cell lines at micro molar concentrations. EDC strongly inhibited the clonogenic propagation of prostate cancer cells. EDC arrests the prostate cancer cells at G2/M phase of the cell cycle and then eventually leads to apoptosis, which was confirmed by DNA fragmentation assay and poly (ADP-ribose) polymerase cleavage western blot analysis. At sub-lethal concentrations, EDC inhibited the migration of PC-3 by 87.1% and DU-145 by 68.0%. Thus, the present study revealed the in vitro cytotoxic and anti-metastatic property of EDC against PC-3 and DU-145 cancer cells.

Andrographolide, a new potential NF-κB inhibitor: docking simulation and evaluation of drug-likeness

Andrographolide is a potent anticancer and anti-inflammatory agent isolated from the plant Andrographis paniculata. It is found to be cytotoxic against various cancer cell lines (in vitro) and also reported to act as an anti-inflammatory agent by interfering with NF-κB protein. Andrographolide induced higher percentage of apoptosis in glutathione-depleted lymphoma cell lines. Andrographolide was also reported to form dehydrated adduct with reduced glutathione at 50° C. On the basis of these observations, the docking analysis of andrographolide with its target protein (NF-κB/p50) and its proposed anti-target protein (glutathione S-transferase) was carried out. Docking analysis predicted the best putative pose of andrographolide molecule in the active site of NF-κB and glutathione S-transferase proteins. Here we report that the furan ring of andrographolide interacts with cysteine 59 of NF-κB/p50 and thereby inhibiting the protein action. Docking studies showed the andrographolide binding to the H-site of glutathione S-transferase enzyme which resembles the behaviour of susceptible xenobiotics inactivated by glutathione S-transferase enzyme. Andrographolide obeys Pfizers rule but drug-likeness value for andrographolide is found to be negative as the molecule showed low drug score. Hence, inactivation by glutathione S-transferase and low drug score could possibly be the reasons to make andrographolide inactive in clinical trial.

Bioassay-guided isolation and evaluation of antiproliferative effects of (Z)-ethylidene-4, 6-dimethoxycoumaran-3-one from Pogostemon Quadrifolius (Benth.)

The aim of the study was to isolate and identify the major cytotoxic principle from plant leaves of Pogostemon quadrifolius (Benth.) and evaluate its antiproliferative potential against human cancer cells. Plant leaves were extracted sequentially with a soxhlet apparatus, using petroleum ether, chloroform and methanol solvents. Petroleum ether and chloroform extracts exhibited antiproliferative properties against Caco-2, HeLa, THP-1, MCF-7 and Jurkat E6-1cancer cell lines tested, but methanol extracts failed to exhibit such activity. The major antiproliferative principle from petroleum ether and chloroform extracts was isolated with the help of bioassay guided column chromatography. This cytotoxic compound was further analysed by UV, TLC, HPLC, LC-MS, GC-MS and NMR analyses and was identified to be novel: (Z)-ethylidene-4,6-dimethoxycoumaran-3-one (Compound 1). The half-maximal inhibitory concentrations for proliferation (IC50) exhibited by compound 1 were 19.4, 23.1, 22.1, 35.9 and 8.32 µM against Caco-2, HeLa, THP-1, MCF-7 and Jurkat E6-1 cancer cell lines, respectively. Further experiments revealed that compound 1 triggered the apoptosis mode of cell death in cancer cell lines. Thus, the present study allowed isolation and identification of a novel cytotoxic natural compound, (Z)-ethylidene-4,6-dimethoxycoumaran-3-one, from plant leaves of P. quadrifolius (Benth.). Our pre-clinical study also indicated that compound 1 is particularly active in the acute T cell leukemia cell line (Jurkat E6-1) with potential for application as a chemotherapeutic agent in the future.

Dimethyl sulfoxide inactivates the anticancer effect of cisplatin against human myelogenous leukemia cell lines in in vitro assays

Objectives: To investigate the effect of DMSO on cisplatin induced cytotoxicity (invitro) against K562 (Human mylogenous leukemia) cell line and to study the cisplatin-DMSO adduct formation using UV-spectrophotometer. Materials and methods: Effect of DMSO on the cytotoxicity of cisplatin was studied in K562 (Chronic mylogenous leukemia) cell line by MTT assay. Cisplatin-DMSO adduct formation was studied by continuously monitoring the increase in absorption peaks for 30 minutes using UV-spectrophotometer. Results: 0.1-0.3% DMSO markedly reduced the cytotoxic activity of cisplatin in K562 cells. Cisplatin-DMSO adduct formation was detected using UV-spectrophotometer. Continuous increase in UV absorbance between 250nm-290nm was observed when cisplatin (0.5mg/ml) and DMSO (10%) were mixed. Conclusion: Present study revealed that DMSO inactivates the cytotoxicity of cisplatin. Cisplatin-DMSO mixture showed increased absorbance at 250-290nm. Therefore, using DMSO in invitro assays might result in misinterpretation of actual efficacy of drugs.