In vitro toxicity to breast cancer cells of microsphere-delivered mitomycin C and its combination with doxorubicin.

To better understand and design microsphere systems for the locoregional delivery of anticancer drug combinations to solid tumors, (1) the cytotoxicity of microsphere-delivered mitomycin C (MMC) was evaluated and (2) various schedules of MMC and doxorubicin (Dox) were tested for their toxicity in vitro towards a murine breast cancer cell-line, EMT6. To accomplish the former MMC was loaded onto oxidized sulfopropyl dextran microspheres, released in a pH 7.4 buffer solution and tested for its potency against EMT6 cells versus a standard MMC solution. For the latter EMT6 cells were exposed to MMC or Dox as single agents or together using various drug concentrations and schedules. The efficacy of the treatments was measured using a clonogenic assay. MMC released from the microspheres showed similar activity against EMT6 cells to freshly prepared MMC solutions. Greater-than-additive toxicity was observed when MMC was given either simultaneously or after Dox exposure. In contrast, administration of MMC before Dox exposure resulted in toxicity that ranged from additive to sub-additive; this reduced toxicity was mainly due to increasing cell density arising from the design of the assay. These results help explain our previous in vivo investigations using microsphere-delivered combinations of the same agents in EMT6 solid tumors.     

In vivo efficacy and toxicity of intratumorally delivered mitomycin C and its combination with doxorubicin using microsphere formulations

The efficacy and toxicity of intratumorally (i.t.) administered anticancer drugs mitomycin C (MMC) and doxorubicin (Dox) incorporated in polymeric microspheres were investigated. Biodegradable sulfopropyl dextran microspheres and their oxidized products were used to load Dox and MMC, respectively. EMT6 mouse mammary cancer cells were injected into the hind leg of BALB/c mice. MMC microspheres, alone or combined with Dox microspheres, were injected i.t. once tumors had reached around 0.3 g. The tumor-plus-leg diameter was measured daily and the delay in time for the tumor to grow to 1.13 g relative to control (TGD) was employed as an indication of therapeutic effect. General toxicity was determined by monitoring weight, appearance and behavior of the mice. Morphology and histology of tumor and heart tissues were also examined. An average 79% TGD was observed after i.t. injection of MMC microspheres. The i.t. co-administration of MMC and Dox microspheres resulted in a 185% TGD. The i.t. injections of the microsphere formulations did not result in visible signs of toxicity in animals. In contrast, systemic (i.e. i.p.) injections of MMC solutions caused considerable general toxicity. This study suggests that i.t. delivery of anticancer drugs by polymeric microspheres is an effective way of improving the therapeutic index for cancer chemotherapy of selected solid tumors under special conditions.     

Liposome-encapsulated doxorubicin reverses drug resistance by inhibiting P-glycoprotein in human cancer cells

The most frequent drawback of doxorubicin is the onset of drug resistance, due to the active efflux through P-glycoprotein (Pgp). Recently formulations of liposome-encapsulated doxorubicin have been approved for the treatment of tumors resistant to conventional anticancer drugs, but the molecular basis of their efficacy is not known. To clarify by which mechanisms the liposome-encapsulated doxorubicin is effective in drug-resistant cancer cells, we analyzed the effects of doxorubicin and doxorubicin-containing anionic liposomal nanoparticles ("Lipodox") on the drug-sensitive human colon cancer HT29 cells and on the drug-resistant HT29-dx cells. Interestingly, we did not detect any difference in drug accumulation and toxicity between free doxorubicin and Lipodox in HT29 cells, but Lipodox was significantly more effective than doxorubicin in HT29-dx cells, which are rich in Pgp. This effect was lost in HT29-dx cells silenced for Pgp and acquired by HT29 cells overexpressing Pgp. Lipodox was less extruded by Pgp than doxorubicin and inhibited the pump activity. This inhibition was due to a double effect: the liposome shell per se altered the composition of rafts in resistant cells and decreased the lipid raft-associated amount of Pgp, and the doxorubicin-loaded liposomes directly impaired transport and ATPase activity of Pgp. The efficacy of Lipodox was not increased by verapamil and cyclosporin A and was underwent interference by colchicine. Binding assays revealed that Lipodox competed with verapamil for binding Pgp and hampered the interaction of colchicine with this transporter. Site-directed mutagenesis experiments demonstrated that glycine 185 is a critical residue for the direct inhibitory effect of Lipodox on Pgp. Our work describes novel properties of liposomal doxorubicin, investigating the molecular bases that make this formulation an inhibitor of Pgp activity and a vehicle particularly indicated against drug-resistant tumors.     

Dose response and toxicity of doxorubicin microspheres in a rat tumor model.

The therapeutic response and toxic effects of chemotherapy using several doses of doxorubicin in conventional solution form or bound to an ion-exchange resin were compared in a rat tumor model, to assess the relationship of drug dose to therapeutic efficacy and associated toxicity. Single bolus injections of 3.0, 4.5, 6.0, 7.5 and 9.0 mg/kg were administered via the abdominal aorta to rats bearing hindlimb tumors. Tumor size was measured serially and the growth rates of treated groups were compared with a control growth curve. In addition, the effect of empty microspheres on tumor growth rate was assessed. The levels of circulating white blood cells were measured and compared to control levels to provide an indication of the severity of bone marrow toxicity experienced by each form of treatment. Finally, any difference in the distribution of doxorubicin to tumor, hindlimb and cardiac tissue following administration of doxorubicin as free drug or on microspheres was ascertained. Empty ion-exchange resin exerted a small although significant detrimental effect on tumor growth which may be explained by the embolization of microspheres in the precapillary blood vessels of the tumor resulting in a transient delay in tumor growth rate. The lowest dose of doxorubicin produced a significantly better therapeutic response when administered in the free drug form, but higher doses elicited an equivalent delay in tumor growth for both drug microsphere and free drug groups in a dose-dependent manner, with the maximum anti-tumor response occurring at the highest dose. Treatment with free doxorubicin at high doses resulted in significant reductions of circulating white blood cells suggesting the occurrence of bone marrow toxicity.(ABSTRACT TRUNCATED AT 250 WORDS)     

In vivo evaluation of a new polymer-lipid hybrid nanoparticle (PLN) formulation of doxorubicin in a murine solid tumor model.

The purpose of this study is to evaluate the in vivo efficacy, unwanted toxicity and loco-regional distribution of a doxorubicin-loaded polymer-lipid hybrid nanoparticle (Dox-PLN) formulation in a murine solid tumor model after intratumoral injection. Dox-PLN were prepared by dispersing Dox in stearic acid and tristearin, with subsequent addition of a novel anionic polymer HPESO (hydrolyzed polymer of epoxidized soybean oil) to enhance the drug incorporation in the lipids. Solid tumors were obtained by injecting EMT6 mouse mammary cancer cells intramuscularly into the hind legs of BALB/c mice. Dox-PLN, blank PLN or surfactant formulations were injected intratumorally (IT) when tumors reached approximately 0.3 g. In vivo efficacy of treatment was measured by tumor growth delay (TGD), defined as the delay in time for the tumor to grow to 1.13 g relative to the untreated control. Signs of unwanted drug toxicity, the histology and morphology of tumor and heart tissues, and the IT distribution of Dox-PLN after IT treatment were examined or monitored. IT-administered Dox-PLN resulted in 70% and 100% TGD (p<0.01) for Dox doses of 0.1 and 0.2 mg, respectively. Dox-PLN treated tumors developed substantially larger central necrotic regions than the untreated tumors, with Dox-PLN residues extensively distributed among the dead cell debris, suggesting that the anticancer effect of Dox-PLN was mainly a combined result of IT nanoparticle distribution and short-ranged, sustained drug release. Except for two of fifteen mice receiving the higher 0.2 mg Dox dose showing transient fur-roughing, all Dox-PLN treated mice showed no signs of toxicity. The present study demonstrates that Dox-PLN possess significant in vivo cytotoxic activity against solid tumors with minimal systemic toxicity. IT administered Dox-PLN have the potential to improve the therapeutic index of loco-regional solid tumor chemotherapy.     

Doxorubicin-loaded casein microspheres: protein nature of drug incorporation.

We have studied incorporation of [14C]doxorubicin within protease-sensitive casein microspheres both by 14C-activity, measuring total drug, and HPLC, measuring free drug only. It was found that total drug content (27.7 micrograms mg-1) exceeded free drug content (3.2 micrograms mg-1) suggesting that the major portion of doxorubicin was incorporated via a covalent linkage to matrix protein. In-vivo drug disposition and activity studies suggested that this fraction of doxorubicin was the major species within tumour tissue (total vs free: 5 min, 14.3 micrograms g-1 vs 0.7 micrograms g-1; 24 h, 11.7 micrograms g-1 vs 1.1 micrograms g-1; 48 h, 11.2 micrograms g-1 vs 1.2 micrograms g-1; 72 h, 10.0 micrograms g-1 vs 0.8 micrograms g-1), did not exhibit a 'burst' effect, was slowly cleared (30% loss over 3 days), and was equiactive (growth delay = 12 days) compared with drug in solution (growth delay = 10 days). This work clearly implicates in-vivo microsphere matrix biodegradation in drug release and subsequent disposition and activity.     

Covalent coupling of doxorubicin in protein microspheres is a major determinant of tumour drug disposition

Doxorubicin is shown to be present in albumin microspheres (10-40 microns) in two forms: the native drug and a fraction of drug covalently coupled to the protein matrix probably via glutaraldehyde. Upon trypsin digestion the fraction covalently coupled is released and can be resolved from native doxorubicin by high performance liquid chromatography and quantitated either by using 14C-labelled doxorubicin or by measuring the absorption of the doxorubicin chromophore at 480 nm. Albumin microspheres contained 6.9 micrograms/mg protein covalently bound drug versus 11.1 micrograms/mg native drug when 1% glutaraldehyde was used in microsphere preparation. The covalently bound fraction increased significantly with 2% glutaraldehyde. Albumin/polyaspartic acid microspheres lacked a covalently bound fraction when prepared under the same conditions as pure albumin microspheres (35 micrograms/mg native drug, 1% glutaraldehyde) but transferrin microspheres contained similar amounts of bound and native albumin. In vivo, albumin microspheres altered the disposition of doxorubicin in a rat mammary carcinoma (Sp107) compared to albumin/polyaspartic acid microspheres by reducing the rate of parent drug elimination from the tumour and by reducing its biotransformation to 7-deoxyaglycone metabolites. These data indicate that covalent coupling is a key component in the way doxorubicin is handled in tumours after administration of protein microspheres.     

维拉帕米对MDR1基因转染的Swiss-3T3细胞的化疗增敏作用

为进一步了解维拉帕米对抗药性逆转作用的特征,在人MDR1基因转染的Swiss-3T3多药抗药性细胞,观察了维拉帕米逆转幅度与阿霉素抗性水平的关系。各个转染细胞与母细胞相比,阿霉素毒性明显降低。非毒性浓度(3μmol·L^-1)的维拉帕米对阿霉素毒性的增强作用,在转染细胞均高于母细胞,但逆转幅度与抗性水平成反比。Southern杂交显示,转染细胞基因组中有MDR1 cDNA整合。转染细胞的阿霉素蓄积障碍可被维拉帕米纠正。讨论了药物主动转运的饱和现象在维拉帕米增强效应中的作用,以及P-糖蛋白与药物相互作用的方式。     

Establishment of doxorubicin-resistant subline derived from HCT15 human colorectal cancer cells

Doxorubicin, one of the clinically most useful anticancer agents, is used alone or in combination with other drugs against a wide variety of tumors, recently. But cancer cells developed resistance to this agent in many ways. This resistance is an important limiting factor of doxorubicin for anticancer drug. We newly established doxorubicin-resistant HCT15/CL02 subline from parental HCT15 human adenocarcinoma colon cancer cells. HCT15/CL02 revealed resistance to doxorubicin about 85-fold of its parental cells, and it also revealed cross-vealed to actinomycin D, etoposide and vinblastine but not to cisplatin and tamoxifen. And verapamil, a reversal agent of multidrug-resistance (MDR) by P-glycoprotein, elevated the cytotoxicity of doxorubicin against both HCT15 and HCT15/CL02 cells. But the relative resistant rate was not reduced. Verapamil had no effects on the toxicity of cisplatin to the both cell lines. These results indicate that HCT15/CL02 cells have some functionally complex mechanisms for MDR.     

Comparison of albumin and casein microspheres as a carrier for doxorubicin

Doxorubicin (Adriamycin)-loaded casein and albumin microspheres, with diameters between 14 and 38 micron (50% weight average) were prepared by glutaraldehyde stabilization of the aqueous phase (containing protein and drug) of a water in oil emulsion. Physical properties, drug loading characteristics and release rates from microspheres in-vitro have been compared and correlated with effects on tumour growth when injected intratumourally in rats. Compared with albumin, the surface charge of the casein system was more negative and the microspheres exhibited a slower release of drug in-vitro. Both observations could be explained by the lower drug content of the casein system. There was evidence for the formation of a doxorubicin complex in the microspheres, the significance of which is not yet known. Casein microspheres containing 11 micrograms of doxorubicin had a similar inhibitory effect on tumour growth (growth delay = 20.7 days) to 85 micrograms of drug incorporated into albumin microspheres (growth delay = 18.6 days). The absence of a simple dose-response relationship shows that carrier matrix can influence potency of incorporated drug. The results are consistent with release rate of the drug from microspheres (obversely, rate of drug delivery to the tumour), being a determinant of potency in these systems.     

Involvement of the multidrug resistance P-glycoprotein in acetaminophen-induced toxicity in hepatoma-derived HepG2 and Hep3B cells

Acetaminophen overdose causes severe hepatic failure. Although the mechanisms of acetaminophen hepatotoxicity have been well investigated, little is known about the involvement of the P-glycoprotein in acetaminophen transport and toxicity. P-Glycoprotein is a membrane efflux pump, playing a significant role in regulating absorption, excretion, and tissue distribution of many drugs. To evaluate the contribution of P-glycoprotein transporter in the course of acetaminophen-induced toxicity, HepG2 and Hep3B cells with different P-glycoprotein expression and activity, were treated by acetaminophen (1-10 mM) for different time periods, with or without the P-glycoprotein inhibitor verapamil. P-Glycoprotein activity was determined by rhodamine 123 efflux assay and western blot analysis. To assess the acetaminophen-induced toxicity and effect of verapamil, we investigated cellular redox status, phosphatidylserine externalization, nuclear fragmentation and ultrastructural changes. Verapamil markedly enhanced acetaminophen-induced oxidative damage and cell death. Moreover, verapamil revealed acetaminophen toxicity even at subtoxic levels. High acetaminophen concentrations increased P-glycoprotein activity and content in both HepG2 and Hep3B cells. These observations suggest the involvement of P-glycoprotein in acetaminophen transport. Notwithstanding the differences of the investigated hepatoma cell lines in P-glycoprotein function, acetaminophen-induced toxicity was similar, possibly due to different functions of drug-metabolizing systems. We conclude that acetaminophen is a P-glycoprotein substrate and P-glycoprotein is involved in acetaminophen transport and toxicity in HepG2 and Hep3B cells. This study establishes the fact that acetaminophen can modulate P-glycoprotein in tumour cells, suggesting that its routine use in cancer patients in combination with anticancer drugs, may influence the result of chemotherapy.     

Drug targeting systems for cancer chemotherapy

Tumour specific drug targeting has been a very actively investigated area for over 2 decades. Various approaches have involved the use of drug delivery systems that can localise the anticancer agent at the tumour site without damaging the normal cells. For this purpose, various delivery systems that have been utilised are liposomes, microspheres and recently, nanoparticles. Two liposome formulations containing anticancer drugs for example, adriamycin and daunomycin are already on the market in the USA and Europe. Microspheres are also being investigated for delivering various anticancer drugs and protein/peptides for anticancer treatment, and several formulations are in Phase I/II clinical trials. Antitumour drugs have also been linked to tumour specific monoclonal antibodies via various chemical linkages. Doxorubicin was linked to a chimeric monoclonal antibody that was targeted to the Lewis Y antigen. Though this conjugate initially showed potential, it was recently dropped from Phase II clinical trials. Another approach with monoclonal antibodies has been the use of immunotoxins. Immunotoxins initially showed promise as potential anticancer agents at picomolar concentrations but several clinical and preclinical studies have not shown much promise in this regard. Drug containing liposomes and microspheres have been further linked to tumour specific monoclonal antibodies to enhance their tumour specificity. Most of the studies with immunoliposomes or targeted microspheres have not gone beyond the preclinical studies. New therapeutic approaches are presently emerging based on natural products like cytokines, peptide growth factor antagonists, antisense oligonucleotides and specific genes. These approaches need the help of delivery systems to deliver these complex molecules to tumour cells. One of the current pursued approaches is the use of cationic liposomes. Several clinical studies are undergoing with various cationic liposomes and the next few years will demonstrate the usefulness of this approach. In recent years, the problems in cancer treatment have been complicated with the emergence of resistance strains leading to resistant and cross-resistant tumour cells. Several agents have been used to overcome or reverse drug-resistance in solid tumours and it remains a highly pursued area in cancer treatment.     

Reversal of acquired resistance to doxorubicin in K562 human leukemia cells by astemizole

This study demonstrates that astemizole, a non-sedating anti-histaminergic drug with low toxicity in vivo, greatly potentiates the growth-inhibitory activity of doxorubicin in doxorubicin-resistant human leukemia cells (K562/DXR). Astemizole synergistically potentiated the cytotoxicity of doxorubicin for K562/DXR cells at a concentration of 0.1-3 microM in a dose-dependent manner, whereas they showed hardly any synthergistic effect in the parental cell line (K562) at the same concentration. Since doxorubicin resistance in these cells is associated with the expression of high levels of P-glycoprotein, we evaluated the effect of astemizole on P-glycoprotein activity in cytofluorographic efflux experiments with doxorubicin. Our results indicate that astemizole inhibits the P-glycoprotein pump-efflux activity in a dose-related manner. Moreover, it also inhibits the photolabeling of P-glycoprotein by [3H]azidopine in a dose-dependent manner. These findings provide a biological basis for the potential therapeutic application of astemizole as an anticancer drug either alone or in combination with doxorubicin to multidrug-resistant leukemic cells.     

Drug targeting systems for cancer chemotherapy

Tumour specific drug targeting has been a very actively investigated area for over 2 decades. Various approaches have involved the use of drug delivery systems that can localise the anticancer agent at the tumour site without damaging the normal cells. For this purpose, various delivery systems that have been utilised are liposomes, microspheres and recently, nanoparticles. Two liposome formulations containing anticancer drugs for example, adriamycin and daunomycin are already on the market in the USA and Europe. Microspheres are also being investigated for delivering various anticancer drugs and protein/peptides for anticancer treatment, and several formulations are in Phase I/II clinical trials. Antitumour drugs have also been linked to tumour specific monoclonal antibodies via various chemical linkages. Doxorubicin was linked to a chimeric monoclonal antibody that was targeted to the Lewis Y antigen. Though this conjugate initially showed potential, it was recently dropped from Phase II clinical trials. Another approach with monoclonal antibodies has been the use of immunotoxins. Immunotoxins initially showed promise as potential anticancer agents at picomolar concentrations but several clinical and preclinical studies have not shown much promise in this regard. Drug containing liposomes and microspheres have been further linked to tumour specific monoclonal antibodies to enhance their tumour specificity. Most of the studies with immunoliposomes or targeted microspheres have not gone beyond the preclinical studies. New therapeutic approaches are presently emerging based on natural products like cytokines, peptide growth factor antagonists, antisense oligonucleotides and specific genes. These approaches need the help of delivery systems to deliver these complex molecules to tumour cells. One of the current pursued approaches is the use of cationic liposomes. Several clinical studies are undergoing with various cationic liposomes and the next few years will demonstrate the usefulness of this approach. In recent years, the problems in cancer treatment have been complicated with the emergence of resistance strains leading to resistant and cross-resistant tumour cells. Several agents have been used to overcome or reverse drug-resistance in solid tumours and it remains a highly pursued area in cancer treatment.     

A cell-based drug delivery system for lung targeting: II. Therapeutic activities on B16-F10 melanoma in mouse lungs

The in vitro and in vivo anticancer activities of doxorubicin-loaded B16-F10 murine melanoma cells (DLTC) were evaluated. DLTC showed similar growth-inhibitory effects against live B16-F10 cells with doxorubicin solution in cell culture system, with the IC₅₀     

Relationship between reductive drug metabolism in tumour tissue of anthracyclines in microspherical form and anti-tumour activity

Increased activity against a rat solid tumour of doxorubicin incorporated into protein microspheres and administered intratumourally was associated with both increased duration of exposure of tumour tissue to native drug and anaerobic bioreduction of doxorubicin to 7-deoxyaglycones, indicating formation of reactive drug intermediates within tumour tissue. To investigate which of these aspects of drug disposition determined activity we have compared the in vivo fate (clearance from and metabolism by tumour tissue) of doxorubicin in microspherical form with the analogue 4'-deoxydoxorubicin and related this to the tumour growth delay recorded for these drugs. Within the dose range 42 to 55 micrograms, growth delay (14-18 days) of doxorubicin in microspherical form was markedly superior to drug in solution, whereas growth delay of 4'-deoxydoxorubicin in microspherical form (4.3-7.2 days) was not greater than drug in solution. Metabolism to 7-deoxyaglycones by tumour tissue was not a prominent feature of either drug when administered in solution. However, in microspherical form both drugs were extensively metabolized (peak concentrations: 3.6 micrograms/g doxorubicin 7-deoxyaglycone; 2.5 micrograms/g 4'-deoxydoxorubicin 7-deoxyaglycone). Native drug concentrations in tumour tissue were similar after administration in microspherical form at 48 hr (doxorubicin 3.8 micrograms/g; 4'-deoxydoxorubicin 3.7 micrograms/g) and 72 hr (doxorubicin 2.4 micrograms/g; 4'-deoxydoxorubicin 2.7 micrograms/g). At both time points, following administration in microspherical form, tumour tissue concentrations of doxorubicin were significantly greater than when drug was administered in solution, whereas no significant differences were observed for 4'-deoxydoxorubicin. The results are inconsistent with the process of anaerobic bioreduction of doxorubicin to 7-deoxyaglycones being an important component of its anti-tumour activity in microspherical form and point to the importance of increased duration of exposure to native drug.     

A Cell-Based Drug Delivery System for Lung Targeting: II. Therapeutic Activities on B16-F10 Melanoma in Mouse Lungs

The in vitro and in vivo anticancer activities of doxorubicin-loaded B16-F10 murine melanoma cells (DLTC) were evaluated. DLTC showed similar growth-inhibitory effects against live B16-F10 cells with doxorubicin solution in cell culture system, with the IC₅₀     

Hydrogel Matrix Entrapping PLGA-Paclitaxel Microspheres: Drug Delivery with Near Zero-Order Release and Implantability Advantages for Malignant Brain Tumour Chemotherapy

Purpose

To develop paclitaxel-delivering PLGA microspheres entrapped in a gel matrix with sustained drug release properties and implantability advantages for local glioma chemotherapy.

Methods

Paclitaxel-loaded PLGA microspheres were fabricated using electrohydrodynamic atomization and entrapped by electrospray and gelation. The physicochemical characterizations were performed using scanning electron microscopy and differential scanning calorimetry. The influence of various parameters on the disintegration time was investigated. In vitro release of paclitaxel was quantified using high performance liquid chromatography. Cytotoxicity of the formulations was assessed by the quantification of IC₅₀ and caspase-3 activity against C6 glioma cells in vitro. The formulations were tested against a subcutaneous C6 glioma tumour in mice.

Results

Highly monodisperse gel beads containing a uniform microsphere distribution were obtained. Gelation using Ca²⁺ ions ensured entrapment of microspheres with high loading efficiency. With an increase in the gelation time, gelling bath concentration and decrease in microsphere loading, it was more difficult to disintegrate the beads and release the microspheres. The formulations demonstrated sustained drug release for more than 60 days at a near-constant rate and a low initial burst. Cell culture studies proved the cytotoxicity against C6 glioma and improved performance in comparison to Taxol®. The formulations could reduce subcutaneous tumour volume to a greater extent compared to Taxol® and the control.

Conclusions

Paclitaxel-loaded PLGA microspheres entrapped in an alginate gel matrix could be potential local chemotherapy implants to treat malignant glioma with critical advantages of implantability and sustained drug release with low initial burst.

     

Mucoadhesive microspheres containing gentamicin sulfate for nasal administration: preparation and in vitro characterization

In this study, suitable microsphere formulations were designed in order to provide the absorption of a high polar drug through nasal mucosa. For this purpose, gentamicin sulfate (GS) was chosen as a model drug and used at different drug/polymer ratios in the microsphere formulations. The microspheres were prepared by spray drying technique. Hydroxypropyl methylcellulose was used as a mucoadhesive polymer in the formulations to increase the residence time of the microspheres on the mucosa. Sodium cholate was added into the formulations for increasing the absorption of GS through nasal mucosa. The in vitro characteristics of the microspheres were determined. The microspheres were evaluated with respect to the particle size, production yield, encapsulation efficiency, shape and surface properties, drug-polymer interaction, mucoadhesive property, in vitro drug release and suitability for nasal drug delivery.     

Antitumour Activity of Mitoxantrone-loaded Chitosan Microspheres Against Ehrlich Ascites Carcinoma

Glutaraldehyde cross-linked chitosan microspheres containing the antineoplastic agent mitoxantrone were prepared and the antitumour activity was evaluated against Ehrlich ascites carcinoma in mice by intraperitoneal injections. The tumour inhibitory effect was followed by monitoring animal survival time and change in body weight for a period of 60 days. While the mean survival time of animals which received 2 mg and 1 mg of free mitoxantrone intraperi-toneally was 2.1 and 4.6 days, respectively, animals which received 2 mg mitoxantrone via microspheres showed a mean survival time of 50 days. Five out of 8 animals treated using microspheres lived beyond 60 days. The percentage ratio mean survival time of the treated group divided by the mean survival time of the untreated group for animals treated using mitoxantrone-loaded chitosan microspheres containing 2 mg of the drug was 290 compared with 12.2 for those which received 2 mg of the free drug. The antitumour effect of mitoxantrone-loaded microspheres against Ehrlich ascites carcinoma was much higher than that of doxorubicin-loaded microspheres reported by previous workers. Our data demonstrate the potential of mitoxantrone-loaded chitosan microspheres for sustained drug delivery to minimize drug toxicity and maximize therapeutic efficacy.