Unravelling the antimetastatic potential of pentoxifylline, a methylxanthine derivative in human MDA-MB-231 breast cancer cells.

Pentoxifylline (PTX), a methylxanthine derivative is a non-steroidal immunomodulating agent with unique hemorheologic properties. It is used in the treatment of intermittent claudication as it increases the amount of oxygen reaching tissues by increasing the flexibility of red blood cells. Recently, it has also shown to exhibit anti-metastatic and anti-angiogenic activities in B16F10 melanoma cells both in vitro as well as in vivo. As per the reports, the choice of drug in the treatment of breast cancer is paclitaxel, but the major limitation is its toxicity. However, the effects of PTX on metastatic processes in breast cancer are not currently known. Therefore, in this study, we have examined the effect of PTX in MDA-MB-231 human breast cancer cells. The MTT assay showed dose- and time-dependent decreases in cellular proliferation. The non-toxic concentration of PTX selected were 1, 2.5 and 5 mM for 24 h. PTX induced a G0-G1 cell-cycle arrest leading to apoptosis. Further, it affected adhesion to both the matrigel and collagen type-IV in a time- and dose-dependent manner. The PTX impeded the migration of MDA-MB-231 cells and also decreased the activities of both MMP-2 and MMP-9. Thus, PTX at non-toxic doses affected cellular proliferation, adhesion, migration and invasion. These results demonstrate its anti-metastatic effect on MDA-MB-231 cells, and further studies need to be carried out to understand the mechanism of action.

Lecithin-based novel cationic nanocarriers (Leciplex) II: improving therapeutic efficacy of quercetin on oral administration.

The objective of the present investigation was to evaluate ability of the novel self-assembled phospholipid- based cationic nanocarriers (LeciPlex) in improving the therapeutic efficacy of a poorly water-soluble natural polyphenolic agent, quercetin (QR), on oral administration. Quercetin loaded LeciPlex (QR-LeciPlex) were successfully fabricated using a biocompatible solvent Transcutol HP. The QR-LeciPlex were characterized for particle size, encapsulation efficiency, zeta potential, and particle morphology by cryo-TEM. UV and fluorescence spectral characterization was carried out to find out the association of QR with LeciPlex. Small angle neutron scattering studies (SANS) were carried out to understand the internal structure of Leciplex and to evaluate the influence of the incorporation of QR in the LeciPlex. Anti-inflammatory and antitumorigenic activity of QR-LeciPlex was determined in comparison to QR suspension to evaluate the potential of LeciPlex in improving oral delivery of QR. QR-LeciPlex exhibited a particle size of ∼400 nm and had excellent colloidal stability. The QR-LeciPlex had a zeta potential greater than +30 mV and exhibited very high encapsulation efficiency of QR (>90%). UV and fluorescence spectral characterization indicated the interaction/association of QR with LeciPlex components. Cryo-TEM studies showed that LeciPlex and QR-LeciPlex have a unilamellar structure. SANS confirmed the unilamellar structure of LeciPlex and indicated that the incorporation of QR does not have any effect on the internal structure of the LeciPlex. QR-LeciPlex exhibited significantly higher anti-inflammatory and antitumorigenic activity (p < 0.01) as compared to that of QR suspension on oral administration.

RP-HPLC Estimation of Ramipril and Telmisartan in Tablets.

A rapid high performance liquid chromatographic method has been developed and validated for the estimation of ramipril and telmisartan simultaneously in combined dosage form. A Genesis C18 column having dimensions of 4.6x250 mm and particle size of 5 mum in isocratic mode, with mobile phase containing a mixture of 0.01 M potassium dihydrogen phosphate buffer (adjusted to pH 3.4 using orthophosphoric acid): methanol:acetonitrile (15:15:70 v/v/v) was used. The mobile phase was pumped at a flow rate of 1.0 ml/min and the eluents were monitored at 210 nm. The selected chromatographic conditions were found to effectively separate ramipril (R(t): 3.68 min) and telmisartan (R(t): 4.98 min) having a resolution of 3.84. The method was validated in terms of linearity, accuracy, precision, specificity, limit of detection and limit of quantitation. Linearity for ramipril and telmisartan were found in the range of 3.5-6.5 mug/ml and 28.0-52.0 mug/ml, respectively. The percentage recoveries for ramipril and telmisartan ranged from 99.09-101.64% and 99.45-100.99%, respectively. The limit of detection and the limit of quantitation for ramipril was found to be 0.5 mug/ml and 1.5 mug/ml respectively and for telmisartan was found to be 1.5 mug/ml and 3.0 mug/ml, respectively. The method was found to be robust and can be successfully used to determine the drug content of marketed formulations.

Amelioration of B16F10 melanoma cells induced oxidative stress in DBA/2 mice by pentoxifylline.

The constantly produced small amounts of reactive oxygen species (ROS) are reduced by anti-oxidant enzymes and cellular scavengers. The oxidative stress developed by defect in ROS clearance can result in cell injury and may lead to carcinogenesis. Pentoxifylline is a methylxanthine derivative with rheologic and membrane modifier property. We have examined whether Pentoxifylline (PTX) ameliorates oxidative stress produced in subcutaneously injected mice with B16F10 melanoma cells. Treatment of mice with PTX significantly reduced oxidative stress and attenuated the altered changes of reduced glutathione and lipid peroxides. Our findings provide an experimental basis for using PTX to attenuate oxidative stress induced by B16F10 melanoma cells in liver and lung of DBA/2 mice.

Effects of niosomal cisplatin and combination of the same with theophylline and with activated macrophages in murine B16F10 melanoma model.

The use of cisplatin is limited due to its certain toxic effects. In the present study niosomes of cisplatin by using span 60 and cholesterol were prepared and investigated for antimetastatic activity in experimental metastatic model of B16F10 melanoma. Theophylline and its combination effect with free cisplatin and niosomal cisplatin were also carried out in the same model. The effect of treatment with activated macrophages alone and in combination with cisplatin, theophylline and niosomal cisplatin was also observed. Treatment with niosomal cisplatin (1 mg/kg) and combination of the same with theophylline (15 mg/kg) showed significant reduction in the number of lung nodules as compared to untreated control as well as with free cisplatin (1 mg/kg). The treatment with activated macrophages (activated by using Muramyl dipeptide) significantly reduced the secondary growth of tumor in lung. Niosomal cisplatin showed a significant protection against weight loss and bone marrow toxicity as compared to free cisplatin. These results suggest that cisplatin encapsulated in niosomes has significant antimetastatic activity and reduced toxicities than that of free cisplatin. However theophylline failed to show antimetastatic effect alone or in combination with cisplatin or with activated macrophages.

Tumor regression of B16F10 melanoma in vivo by prevention of neovascularization: study on theophylline.

The aim of this study was to determine the effect of theophylline on neovascularization and tumor regression in murine B16F10 melanoma. Theophylline had no direct toxicity to host and significantly reduced (p < 0.001) tumor volume and neovascularization in B16F10 melanoma implanted murine model. The effect of theophylline on neovascularization was observed distinctly in histologic analysis. This effect is mediated, in part by blocking endothelial cell proliferation, thereby preventing neovascularization of the tumor. Further investigations with theophylline can elucidate the exact mechanism of action which characterize neovascularization activity.

Inhibition of endothelial cell proliferation and tumor-induced angiogenesis by pentoxifylline.

PURPOSE: In this study we investigated the effect of pentoxifylline (PTX) on tumor-induced neovascularization as well as on different steps involved in the angiogenic process. METHODS: To assess angiogenesis inhibition. we injected intradermally (i.d.) 10 B16-F10 melanoma cells into C57BL/6J mice which were subsequently intraperitoneally (i.p.) inoculated with PTX or saline. On day 7 the number of blood vessels converging to the remnant of injected material was counted and the volumes of incipient tumors were calculated in each case. In vitro growth inhibition by PTX was evaluated in two different cell lines of endothelial origin and in human umbilical vein endothelial cells. Motility assays, as well as zymographic assays carried out to analyze gelatinolytic metalloproteinases and urokinase-type plasminogen activator, were performed in one of the endothelial cell lines. RESULTS: A significant inhibition of tumor-induced angiogenesis was observed in C57B1/6 mice i.p. inoculated with PTX, that paralleled reduced incipient tumor volumes. The endothelial cells derived from different sources were inhibited in a dose-response manner by PTX in vitro. Non-cytotoxic PTX concentrations assayed in one of the endothelial cell lines did not inhibit its in vitro cell motility nor its gelatinase secretion, but its low molecular weight urokinase-type plasminogen activator expression. CONCLUSIONS: Our findings suggest that the inhibitory effect of PTX on tumor angiogenesis is related to antiproliferative action on endothelial cells, as well as to down regulation of u-PA secreted by them.

Effect of Caffeine, a xanthine derivative, in the inhibition of experimental lung metastasis induced by B16F10 melanoma cells.

Caffeine, a methyl xanthine derivative, was studied to assess the effect on B16F10 melanoma induced experimental metastasis. Caffeine was administered at a dose of 100 and 50 mg/kg body weight by both routes, to tumour bearing animals. Solid tumour reduction studies with Caffeine showed a significant reduction in tumour volume for 100 mg/kg dose by both oral and i.p. routes. The Caffeine treated metastatic tumour bearing animals significantly (p<0.001) inhibited lung tumour nodules. Serum sialic acid levels and lung hydroxyproline contents in the treated groups were significantly (p<0.001) low, when compared with the untreated control animals. In the present study, our results suggest that Caffeine inhibits solid tumour development and pulmonary experimental metastasis induced by B16F10 melanoma cells, in murine model.

Differential anthracycline sensitivity in two related human colon carcinoma cell lines expressing similar levels of P-glycoprotein.

Chemosensitivity of the human colon carcinoma HCT-15 cell line to 4'-epidoxorubicin proved to be 100-fold higher than that of its variant HCT-15 EDR. Confocal scanning microscopy showed significant less drug accumulation in HCT-15 EDR. A 2-fold increase in hsp27 expression was found in HCT-15 EDR, with no alteration in hsp70. The expression of the drug exporter Pgp was similar in both cell lines, despite the lower drug accumulation shown by HCT-15 EDR in respect to HCT-15. Other molecules implicated in the acquisition of enhanced chemoresistance or a more active Pgp variant present in HCT-15 EDR, could explain the phenomenon.

Antimetastatic efficacy of niosomal pentoxifylline and its combination with activated macrophages in murine B16F10 melanoma model.

The aims of the present study were a) to enhance the effectiveness of antimetastatic agent, Pentoxifylline (PTX) by encapsulation in niosomes and b) to investigate the anticancer activity by combination therapy involving activated macrophages and PTX solution/PTX niosomes. Niosomes were prepared by lipid film hydration method. Particle size distribution revealed bimodal distribution with median vesicle size of 462 nm. The entrapment efficacy of PTX niosomes was found to be 9.64%. A cumulative release of 82.43% from niosomal suspension was observed at the end of 21 hours. Intravenous administration of niosomal PTX (6 mg/kg and 10 mg/kg) resulted in significant reduction in lung nodules in an experimental metastatic B16F10 model suggesting accumulation of PTX in a distant target organ-lung. Light microscopic observations of histologic sections showed a decrease in number of tumor islands in the lung. Macrophages activated by intraperitoneal injection of Iscove's Modified Dulbecco's Medium (IMDM) containing 20% fetal calf serum (FCS) followed by in vitro incubation with muramyl dipeptide (MDP) were more effective in controlling tumor spread than those activated by FCS alone. Combination therapy of activated macrophages and PTX solution/niosomal PTX showed no additive or synergistic effect in controlling tumor spread. Carbon clearance studies revealed that PTX inhibits the phagocytic ability of activated macrophages, thereby resulting in the failure of combination therapy.

Enhancement of anti-metastatic activity of pentoxifylline by encapsulation in conventional liposomes and sterically stabilized liposomes in murine experimental B16F10 melanoma model.

Pentoxifylline has been shown to exhibit anti-metastatic activity by inhibiting homing of B16F10 melanoma cells in the murine experimental metastasis model. In this study, the effect of encapsulation of pentoxifylline in conventional and sterically stabilized liposomes on its anti-metastatic activity in the murine experimental metastasis model was investigated. After a single intravenous dose (10, 20 or 40 mg kg(-1)), pentoxifylline solution, as well as conventional pentoxifylline liposomes, significantly reduced the number of pulmonary nodules compared with the untreated control group. Conventional pentoxifylline liposomes showed significantly higher inhibition (69%) of pulmonary tumour nodule formation at a dose of 20mg kg(-1) as compared with pentoxifylline solution (49%) at the same dose. Encapsulation of pentoxifylline in sterically stabilized liposomes prepared by incorporation of monomethoxypolyethyleneglycol (5000)-cholesteryl ester further enhanced the inhibition of pulmonary nodule formation (77%) at a dose of 20 mg kg(-1) as compared with conventional pentoxifylline liposomes. Overall, the results suggest that encapsulation of pentoxifylline in conventional liposomes enhanced its anti-metastatic activity. Steric stabilization of pentoxifylline liposomes also resulted in a two-fold increase in anti-metastatic activity (at dose of 10 mg kg(-1)) as compared with conventional liposomes.

Studies on the mechanisms responsible for inhibition of experimental metastasis of B16-F10 murine melanoma by pentoxifylline.

Pentoxifylline (PTX), a methylxanthine derivative widely used as a hemorheological agent in the treatment of peripheral vascular disease, was studied to unveil the mechanisms responsible for its inhibitory action on B16-F10 experimental metastasis. In vitro pretreatment of B16-F10 cells with noncytotoxic concentrations of PTX significantly inhibited their adhesion to reconstituted basement membrane Matrigel(R) and type IV collagen as well as the relative activity of secreted 92 kD metalloproteinase. However, PTX pretreatment of B16-F10 cells did not affect their in vitro invasiveness. Heterotypic organ adhesion assays carried out with B16-F10 cells and suspended organ tissues demonstrated that pretreatment with noncytotoxic concentrations of PTX of both, tumor cells or lung tissue, brought about a dose-dependent inhibition of melanoma cell adhesion to lung. Immunohistochemical studies using antibodies against CD31 adhesion molecule (PECAM-1) revealed that B16-F10 cells adhere to lung endothelial cells. Our results suggest that PTX may exert its inhibitory effect on tumor lodgment, and as a consequence of that on experimental metastases, through an inhibitory action on cell adhesion molecules.

Inhibition of lung homing of B16F10 by pentoxifylline, a microfilament depolymerizing agent.

B16 melanoma has proved to be an ideal model for investigating metastasis. The parental B16F1 line grows as a localized subcutaneous tumor in C57BL/6 or DBA/2 mice. However, by means of successive intravenous transplantation, a subline B16F10 has been established. This shows preponderant lung homing when transplanted by intravenous route into C57BL/6 mice. In this paper we have shown that pentoxifylline (PTX; Hoechst), a microfilament depolymerising agent can inhibit significantly this lung homing of B16F10 cells. It also had a marginal inhibitory effect on subcutaneous tumor growth.

Effect of pentoxifylline, a multidrug resistance reversal agent, on haemopoietic stem cell homing.

This paper investigates the mechanism of mouse haemopoietic stem cell homing into the cytoskeleton depolymerizing agent Pentoxifylline was used, and shown to inhibit stem cell homing. The inhibition was reversible after 6 hours. The results obtained suggest that the haemopoietic stem cell homing receptor is anchored to cytoskeletal support intracellularly.

Survey of services to patients in general practice.

Family doctors vary in the range of services they provide for their patients. Of 267 practices in Devon and Cornwall, 245 responded to a questionnaire sent in September 1989 asking for information about services to patients. Most doctors consulted at six to eight patients per hour, whether or not an appointment system operated. About two-thirds of patients had access to a female GP. Nine out of ten practices employed a practice nurse. Almost all offered a surgery on Saturday mornings. Most surgeries took the phone over at 8.30 a.m. in the week and started consulting at 9 a.m. Almost half were consulting until 6.30 p.m. or later on at least one day per week. 80% of practices were offering a non-urgent appointment on the same or next day. All practices offered childhood immunisation. 80% offered some form of personal list system. 80% offered minor operations, one-third manipulations, 10% homeopathy, 6% hypnosis and 5% acupuncture.

Effect of Tween 80 on adriamycin cytotoxicity in murine P388 leukemia.

Studies on mice bearing P388 adriamycin sensitive and resistant lymphocytic leukemia were carried out to compare antitumor activity of adriamycin dissolved in Tween 80 (10% in distilled water) with that of adriamycin dissolved in distilled water alone. The antitumor activity of adriamycin dissolved in acqueous solutions of Tween 80 was higher than that of adriamycin dissolved in distilled water against P388 leukemia sensitive to adriamycin. However, no change in the activity was observed in P388 leukemia resistant to adriamycin when the drug was administered either in aqueous solution of Tween 80 or in distilled water alone.

Induced resistance in leukaemia L1210 to adriamycin and its cross-resistance to vincristine and bouvardin.

The induction of complete resistance to adriamycin in L1210 leukemia was accomplished after 10 transplant generations. The adriamycin-resistant subline showed cross-resistance to vincristine and bouvardin. It was sensitive to methotrexate; this was observed by a 50% increase in life span compared to the life span of untreated control animals.

Myasthenia gravis in Melanesians in the Solomon islands.

Myasthenia gravis is a rare neuromuscular disease believed to be due to an autoimmune reaction affecting the neuromuscular junction. Two cases occurring in Melanesians of the Solomon Islands are described. Standard treatment includes anticholinesterase medication, and corticosteroids may be required for severe disease. Thymectomy, and plasmapharesis together with immunosuppressive therapy are other measures that may be considered in severe, refractory or generalised disease. Treatment is unlikely to be satisfactory in this Melanesian society where patients are irregular in taking medication and in attending hospital.