dolastatin-15 and Colonic-Neoplasms

Phosphoinositide 3-kinase (PI3-K) is an important regulator of oncogenesis and apoptosis in various types of cancers including colon cancer. A combinatorial strategy of using Cyclooxygenase-2 inhibitor, Celecoxib and Dolastatin, a linear peptide from marine mollusks of Indian Ocean origin has shown anti-neoplastic effects in colon cancer in a rat model.. The signal transduction pathway of PI3-K/AKT and the downstream signaling proteins had been studied in an early stage of colon carcinogenesis (DMH induced) by gene and protein expression, apoptotic studies by colonocyte apoptotic bleb assay, intracellular calcium level by fluorescence spectrometry, mitochondrial membrane potential by Rhodamine 123 flow cytometry and Reactive oxygen species measurement. Molecular docking analysis was employed to study the interaction of oncogenic proteins and the ligand, Celecoxib and Dolastatin.. Apoptotic cell index was lowered with DMH while both the drugs increased it and inhibited PI3-K and AKT expression. Docking studies revealed both the proteins targeted by the drugs via an ATP binding site. An increased expression of GSK-3β, pro-apoptotic protein Bad, transcription factor Egr-1, tumor suppressor protein PTEN while a downregulation of G1-associated cell cycle protein, Cyclin D1 and increased intracellular calcium as well as reactive oxygen species were observed. Also, the number of cells having a higher mitochondrial membrane potential was lowered.. Celecoxib and Dolastatin inhibited the tumor development through regulation of the PI3-K/AKT pathway which can act as a novel target for these drugs.. The anti-cancer properties of Dolastatin, a peptide isolated from marine mollusks in colorectal cancer is shown.

Topics: Adenosine Triphosphate; Animals; Apoptosis; Binding Sites; Celecoxib; Cell Cycle Proteins; Colonic Neoplasms; Cyclooxygenase 2 Inhibitors; Depsipeptides; Down-Regulation; Male; Membrane Potential, Mitochondrial; Oxidative Stress; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; Pyrazoles; Rats; Rats, Sprague-Dawley; Reactive Oxygen Species; Signal Transduction; Sulfonamides

Programmed cell death, also known as apoptosis, is an active process occurring in eukaryotic cells and it depends on various sets of pro and anti-apoptotic proteins. Chemoprevention of colorectal cancer can be achieved by inducing apoptosis using synthetic compound, Celecoxib and natural peptide, Dolastatin 15 in an effective manner. But the apoptotic signaling by these two drugs remain unclear. The present study was thus focused on the role of Bcl2 family of proteins and their interplay with p53 in rats during the chemoprevention by these two drugs. After treatment for 6 wk with 1, 2-dimethylhydrazine (DMH), animals showed a marked occurrence of multiple plaque lesions. However, a simultaneous treatment with Celecoxib and Dolastatin 15 decreases such number to a significant level. DMH treatment also decreases the number of apoptotic cells in the colonic enterocytes which were corrected to the normal level by Celecoxib and Dolastatin 15. An increased expression of Bcl2 while other proteins like Bax, Apaf-1, cyt c, and caspases in the apoptotic pathway, and the tumor suppressor proteins, p53 and p21 get down-regulated after DMH treatment which were reverted back to normal with Celecoxib and Dolastatin 15. Also, cells having high mitochondrial membrane potential had been seen to increase to significant levels which were reduced after the administration of these anti-inflammatory drugs. In silico molecular docking studies also showed that Dolastatin 15 and Celecoxib may bind to the active site pocket of Bcl2 , thus revealing the direct target of Dolastatin 15 and Celecoxib apart from binding to COX-2.

Topics: Animals; Anticarcinogenic Agents; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Carcinogenesis; Caspases; Celecoxib; Colon; Colonic Neoplasms; Cyclooxygenase 2 Inhibitors; Depsipeptides; Male; Membrane Potential, Mitochondrial; Mitochondria; Molecular Docking Simulation; Mollusca; Proto-Oncogene Proteins c-bcl-2; Pyrazoles; Rats; Rats, Sprague-Dawley; Signal Transduction; Sulfonamides; Tumor Suppressor Protein p53; Up-Regulation

The marine ecosystem is a unique and enormously rich source of natural products with potential chemopreventive applications in cancer. In the present study, we explored the chemopreventive role and the molecular mechanism of Dolastatin, a linear peptide from an Indian Ocean mollusk, and Celecoxib, a well-established cyclooxygenase-2 (COX-2) inhibitor in an individual as well as in a combination regimen in 1,2-dimethylhydrazine dihydrochloride (DMH)-induced colon carcinogenesis in a rat model. After a 6-week treatment with DMH, morphological analysis revealed a marked occurrence of preneoplastic features in the colonic mucosa, whereas histologically well-characterized dysplasia and hyperplasia were observed in DMH-treated animals. Simultaneous administration of Celecoxib and Dolastatin reduced these features significantly. DMH treatment affected the number of apoptotic cells in colonic enterocytes, which reverted to the normal level with the use of Celecoxib and Dolastatin. Inflammation remains the dominant molecular mechanism in the development of multiple plaque lesions, the carcinogenic lesions in a DMH-induced process that may be mediated by COX-2. Western blot and immunofluorescence analysis revealed a higher expression of COX-2 and nuclear factor-κB, the transcription factors responsible for proinflammatory proteins such as TNFα, and also the inducible nitric oxide synthase in the DMH group, which was further recovered significantly with the use of Celecoxib and Dolastatin. In-silico molecular docking analysis of Dolastatin as a ligand with various regulatory proteins suggests that although the peptide failed to dock to COX-2, it successfully did so with inducible nitric oxide synthase, thereby indicating the potential of this inflammatory protein as a molecular anticancer target in colon carcinogenesis.

Topics: Animals; Apoptosis; Blotting, Western; Celecoxib; Colonic Neoplasms; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Depsipeptides; Fluorescent Antibody Technique; Intestinal Mucosa; Male; Molecular Docking Simulation; NF-kappa B; Nitric Oxide; Nitric Oxide Synthase Type II; Precancerous Conditions; Pyrazoles; Rats; Rats, Sprague-Dawley; Sulfonamides

Dolastatins 10 and 15 are small peptides isolated from the marine sea hare Dolabella auricularia that have been shown to interact with tubulin. Their growth-inhibitory properties were compared using panels of human ovarian and colon-carcinoma cell lines. Both agents were very potent inhibitors of cell growth, with dolastatin 10 being an average of 9.1-fold more potent than dolastatin 15 [mean 50% inhibitory concentrations (IC50 values) 2.3 x 10(-10) and 2.1 x 10(-9) M, respectively; P < 0.05] and more potent than paclitaxel or vinblastine. While neither dolastatin exhibited marked cross-resistance in cisplatin- or etoposide-resistant cell lines, contrasting effects were observed using an acquired doxorubicin-resistant (CH1doxR, 100-fold resistant, P-glycoprotein overexpressing) cell line. Resistance was significantly higher to dolastatin 15 (12.7-fold) than to dolastatin 10 (only 3.2-fold; P < 0.05) and was reversible in both cases by verapamil. In vivo, using a s.c. advanced-stage human ovarian carcinoma xenograft and equitoxic doses, greater activity was observed with dolastatin 10 (6.1-day growth delay) versus 0.4 days for dolastatin 15. A radioimmunoassay for dolastatin 10 (limit of detection in mouse plasma 5 ng/ml) was developed. The rabbit antiserum aslo cross-reacted by 65% with dolastatin 15. Comparative mouse pharmacokinetics following i.v. administration of 1 mg/kg showed that both compounds are rapidly eliminated, but with a shorter second-phase half-life (t1/2 beta) being observed for dolastatin 15 (being detectable for only up to 4 h post-administration), the t1/2 beta being 3 times longer for dolastatin 10. In addition, areas under the plasma concentration-time curve (AUC values) were 1.6-fold higher for dolastatin 10 (333 versus 208 ng ml-1 h). Plasma binding of dolastatin 10 exceeded 90%. The highly sensitive RIA will be useful for pharmacokinetic studies in conjunction with the planned phase I clinical trials of these novel, extremely potent, tubulin-binding agents, of which dolastatin 10 appears to possess the more promising preclinical features.

Topics: Animals; Antineoplastic Agents; Carcinoma; Cell Division; Colonic Neoplasms; Cross Reactions; Depsipeptides; Drug Resistance, Multiple; Female; Half-Life; Humans; Injections, Intravenous; Mice; Mice, Inbred BALB C; Neoplasm Transplantation; Oligopeptides; Ovarian Neoplasms; Rabbits; Radioimmunoassay; Random Allocation; Transplantation, Heterologous; Tumor Cells, Cultured; Verapamil