Showing papers on "Doxorubicin published in 1994"

Prolonged circulation time and enhanced accumulation in malignant exudates of doxorubicin encapsulated in polyethylene-glycol coated liposomes.

Abstract: In preclinical studies, a doxorubicin liposome formulation containing polyethylene-glycol (Doxil) shows a long circulation time in plasma, enhanced accumulation in murine tumors, and a superior therapeutic activity over free (unencapsulated) doxorubicin (DOX). The purpose of this study was to characterize the pharmacokinetics of Doxil in cancer patients in comparison with free DOX and examine its accumulation in malignant effusions. The pharmacokinetics of doxorubicin and/or liposome-associated doxorubicin were analyzed in seven patients after injections of equivalent doses of free DOX and Doxil and in an additional group of nine patients after injection of Doxil only. Two dose levels were examined, 25 and 50 mg/m2. When possible, drug levels were also measured in malignant effusions. The plasma elimination of Doxil followed a biexponential curve with half-lives of 2 and 45 h (median values), most of the dose being cleared from plasma under the longer half-life. Nearly 100% of the drug detected in plasma after Doxil injection was in liposome-encapsulated form. A slow plasma clearance (0.1 liter/h for Doxil versus 45 liters/h for free DOX) and a small volume of distribution (4 liters for Doxil versus 254 liters for free DOX) are characteristic of Doxil. Doxorubicin metabolites were detected in the urine of Doxil-treated patients with a pattern similar to that reported for free DOX, although the overall urinary excretion of drug and metabolites was significantly reduced. Doxil treatment resulted in a 4- to 16-fold enhancement of drug levels in malignant effusions, peaking between 3 to 7 days after injection. Stomatitis related to Doxil occurred in 5 of 15 evaluable patients and appears to be the most significant side effect in heavily pretreated patients. The results of this study are consistent with preclinical findings indicating that the pharmacokinetics of doxorubicin are drastically altered using Doxil and follow a pattern dictated by the liposome carrier. The enhanced drug accumulation in malignant effusions is apparently related to liposome longevity in circulation. Further clinical investigation is needed to establish the relevance of these findings with regard to the ability of liposomes to modify the delivery of doxorubicin to solid tumors and its pattern of antitumor activity.

The human multidrug resistance-associated protein MRP is a plasma membrane drug-efflux pump.

Abstract: The multidrug-resistance associated protein MRP is a 180- to 195-kDa membrane protein associated with resistance of human tumor cells to cytotoxic drugs. We have investigated how MRP confers drug resistance in SW-1573 human lung carcinoma cells by generating a subline stably transfected with an expression vector containing MRP cDNA. MRP-overexpressing SW-1573 cells are resistant to doxorubicin, daunorubicin, vincristine, VP-16, colchicine, and rhodamine 123, but not to 4'-(9-acridinylamino)methanesulfon-m-anisidide or taxol. The intracellular accumulation of drug (daunorubicin, vincristine, and VP-16) is decreased and the efflux of drug (daunorubicin) is increased in the transfectant. The decreased accumulation of daunorubicin is abolished by permeabilization of the plasma membrane with digitonin, showing that MRP can lower the intracellular daunorubicin level against a concentration gradient. Anti-MRP antisera predominantly stain the plasma membrane of MRP-overexpressing cells. We conclude that MRP is a plasma membrane drug-efflux pump.

Interference by doxorubicin with DNA unwinding in MCF-7 breast tumor cells

Abstract: The capacity of doxorubicin to inhibit topoisomerase II in the MCF-7 breast tumor cell line is supported by the induction of protein-associated single-strand breaks in DNA, as well as by interference with the decatenation activity of nuclear extracts. Doxorubicin also produces non-protein-associated DNA strand breaks (at a supraclinical concentration of 5 microM), which may indicate damage mediated via the generation of free radicals. However, no strand breaks are detected in DNA of MCF-7 cells at the IC50 for doxorubicin (approximately 0.1 microM). At doxorubicin concentrations of 0.05, 0.1, and 0.5 microM, at which growth is inhibited by approximately 15, 50, and 75%, respectively, doxorubicin interferes with radiation-induced unwinding of DNA; doxorubicin also produces a concentration-dependent inhibition of DNA synthesis that corresponds closely to growth inhibition. These studies suggest that DNA strand breaks fail to fully account for the antiproliferative activity of doxorubicin in the MCF-7 breast tumor cell line. Compromised DNA synthesis associated with interference with DNA unwinding may contribute to growth inhibition in MCF-7 cells exposed to doxorubicin.

Anthracycline antibiotics in cancer therapy. Focus on drug resistance

Abstract: 30 years ago an anthracycline antibiotic was shown to have antineoplastic activity. This led to the development of well over 1000 analogues with a vast spectrum of biochemical characteristics. Many biological actions have been described. The original anthracyclines are active against many types of cancer and are an integral part of several curative combinations. They are ineffective against other tumours. Although some analogues show an altered spectrum of activity or an improved therapeutic index relative to the older agents, it is not clear that cardiotoxicity can be totally avoided with these agents. Primary and secondary resistance to anthracyclines remain major clinical problems. Pharmacokinetic studies have been of limited help in explaining this. Overexpression of a surface-membrane permeability glycoprotein (Pgp) was identified in ovarian cancer of patients who had clinical multidrug resistance in 1985. This led the way for the discovery of a number of resistance mechanisms in vitro. Some of these have been found in more than 1 type of cell line, and more than 1 mechanism may exist in a single cell. Additional resistance proteins have been identified, qualitative and quantitative alterations of topoisomerase II have been described, and some mechanisms in other systems have not yet been identified. Some of these may prove to be important in clinical drug resistance. Drugs such as calcium antagonists and cyclosporin, studied initially for their ability to block the Pgp pump, appear to be heterogeneous in this capacity and may have additional sites of action. It will be critical for clinical studies to define the precise resistance mechanism(s) that must be reversed. To date this has been difficult, even in trials ostensibly dealing with the original Pgp. Liposomes can potentially alter toxicity and target drug delivery to specific sites. In addition, they may permit the use of lipophilic drugs that would otherwise be difficult to administer systemically. Resistant tumours may be sensitive to anthracyclines delivered by liposomes. To reduce cardiac toxicity, administering doxorubicin (adriamycin) by slow infusion through a central-venous line should be considered whenever feasible. Monitoring of cardiac ejection fraction and the use of endomyocardial biopsy will permit patients to be treated safely after they reach the dose threshold at which heart failure begins to be a potential risk. A number of structurally modified anthracyclines with the potential advantages of decreased cardiotoxicity and avoidance of multidrug resistance mechanisms are entering clinical trials. Meanwhile, the vast weight of clinical experience leaves doxorubicin as a well tolerated and effective choicc for most potentially anthracycline-sensitive tumours.

Tumour tropism and anti-cancer efficacy of polymer-based doxorubicin prodrugs in the treatment of subcutaneous murine B16F10 melanoma.

Abstract: Doxorubicin (5 mg kg-1) was administered intravenously to C57 mice bearing subcutaneous B16F10 melanomas, distributing into the tumour with an area under the concentration-time curve (0-48 h; AUC) of 8.7 micrograms h g-1. Injection of doxorubicin-N-(2-hydroxypropyl)methacrylamide (HPMA) copolymer conjugate, containing 5 mg of doxorubicin equivalent per kg, mediated an AUC for free doxorubicin (i.e. doxorubicin released from the conjugate) of 15.2 micrograms h g-1 and for total doxorubicin (i.e. free plus conjugated) of 149.1 micrograms h g-1. An increased dose of doxorubicin-HPMA copolymer conjugate (18 mg of doxorubicin equivalent per kg) produced AUC values of 40.1 micrograms h g-1 and 671.7 micrograms h g-1 for free and total doxorubicin respectively. Hence administration of doxorubicin-HPMA copolymer conjugate achieved rises of 1.7- to 4.6-fold in tumour AUC (free doxorubicin) and 17.19 to 77.0-fold in tumour AUC (total doxorubicin). HPMA copolymers bearing fluorescein isothiocyanate accumulated in vascularised stromal regions, particularly in new growth sites at the tumour periphery. Treatment of mice with doxorubicin-HPMA copolymer conjugate achieved treated/control lifespans up to 320% (three doses of 27 mg of doxorubicin equivalent per kg) compared with only 133% using aggressive regimens of free doxorubicin (3 x 5 mg kg-1).

Modulation of tumor cell response to chemotherapy by the organ environment

Abstract: The outcome of cancer metastasis depends on the interaction of metastatic cells with various host factors. The implantation of human cancer cells into anatomically correct (orthotopic) sites in nude mice can be used to ascertain their metastatic potential. While it is clear that vascularity and local immunity can retard or facilitate tumor growth, we have found that the organ environment also influences tumor cell functions such as production of degradative enzymes. The organ microenvironment can also influence the response of metastases to chemotherapy. It is not uncommon to observe the regression of cancer metastases in one organ and their continued growth in other sites after systemic chemotherapy. We demonstrated this effect in a series of experiments using a murine fibrosarcoma, a murine colon carcinoma, and a human colon carcinoma. The tumor cells were implanted subcutaneously or into different visceral organs. Subcutaneous tumors were sensitive to doxorubicin (DXR), whereas lung or liver metastases were not. In contrast, sensitivity to 5-FU did not differ between these sites of growth. The differences in response to DXR between s.c. tumors (sensitive) and lung or liver tumors (resistant) were not due to variations in DXR potency or DXR distribution. The expression of the multidrug resistance-associated P-glycoprotein as determined by flow cytometric analysis of tumor cells harvested from lesions in different organs correlated inversely with their sensitivity to DXR: increased P-glycoprotein was associated with overexpression ofmdr1 mRNA. However, the organ-specific mechanism for upregulatingmdr1 and P-glycoprotein has yet to be elucidated.

Tissue distribution and therapeutic effect of intravenous free or encapsulated liposomal doxorubicin on human prostate carcinoma xenografts.

Abstract: Background. The authors compared the therapeutic effects of doxorubicin in two formulations: free in saline suspension and encapsulated in sterically stabilized liposomes composed of hydrogenated soy phosphatidylcholine/2cholesterol/polyethylene glycol-distearoyl-phosphatidyl-ethanolamine (Doxil, Liposome Technology, Inc., Menlo Park, CA). Results. Laser scan microscope and microfluoro-meter studies showed that the liposome-encapsulated drug entered the liver, the kidneys, and the tumor in greater quantity and remained in the liver and in the tumor longer than the free drug. The liposome formulation produced a 25-fold increase in doxorubicin at the disease site. Doxil was significantly more effective than the free drug in inhibiting growth and in effecting cures and had only minor and temporary systemic toxic effects. Conclusions. The current study demonstrated the therapeutic efficacy of doxorubicin, encapsulated in sterically stabilized liposomes, against prostate carcinoma. Decreased systemic elimination, increased penetration into the tumor, and long liposome presence with slow drug release into the tumor probably accounted for the enhanced therapeutic effect of doxorubicin in sterically stabilized liposomes. Cancer 1994; 73:1478–84. Method. The drug formulations were injected intravenously to treat human prostate carcinoma PC-3, implanted subcutaneously into nude Swiss mice. Confocal laser scan microscopy and microfluorometry were used to determine tissue distribution and to quantitate drug uptake.

Tumor-selective Prodrug Activation by Fusion Protein-mediated Catalysis

Abstract: A two component system, consisting of a fusion protein and an appropriate prodrug, suited to perform selective tumor therapy in vivo is presented. The fusion protein, due to its humanized carcinoembryonic antigen-specific variable region, specifically binds to carcinoembryonic antigen-expressing tumors and has an enzymatic activity comparable to that of human β-glucuronidase. The prodrug is a nontoxic glucuronide-spacer derivative of doxorubicin decomposing to doxorubicin by enzymatic deglucuronidation. In vivo studies in nude mice bearing human carcinoembryonic antigen-expressing tumor xenografts revealed that 7 days after injection of 20 mg/kg fusion protein a high specificity ratio (>100:1) was obtained between tumor and plasma or tumor and normal tissues. Injection of 250 mg/kg of prodrug at day 7 resulted in tumor therapeutic effects superior to those of conventional chemotherapy without any detectable toxicity. These superior therapeutic effects which were observed using established human tumor xenografts can be explained by the approximately 4–12-fold higher doxorubicin concentrations found in tumors of mice treated with fusion protein and prodrug than in those treated with the maximal tolerable dose of drug alone. The nondetectable toxicity in the animals treated with fusion protein and prodrug is probably caused by up to 5-fold lower drug concentrations in normal tissues compared to the animals treated with doxorubicin. Thus, a more tumor-selective therapy, resulting in stronger therapeutic effects and reduced toxicity seems to be possible by the appropriate use of the humanized nontoxic fusion protein and the nontoxic prodrug.

Pharmacokinetics, Biodistribution and Therapeutic Efficacy of Doxorubicin Encapsulated in Stealth® Liposomes (Doxil®)

Abstract: Doxil® (Stealth® liposomal doxorubicin HCl) Injection is doxorubicin HCl incorporated into long circulating liposomes that contain surface-grafted polyoxyethylene chains. These surface-grafted poly...

Clinical studies of liposome-encapsulated doxorubicin

Abstract: Initial clinical studies with doxorubicin entrapped in the bilayer of phosphatidylglycerol-rich liposomes were hindered by the avid reticuloendothelial system (RES) uptake and by drug leakage from circulating liposomes. In contrast, recent tests of a doxorubicin formulation of polyethyleneglycol-coated liposomes (Doxil) in cancer patients indicate that the drug pharmacokinetic properties are significantly altered, with a prolonged distribution half-life of approximately 2 days. Plasma fractionation studies show that nearly all the drug measured in plasma is in liposome-encapsulated form. The dose of Doxil has been escalated from 25 to 60 mg/m2. Stomatitis is the most significant toxicity, and skin toxicity, in the form of hand-foot syndrome, may complicate the repeated administration of Doxil. A number of objective antitumor responses in a variety of malignancies have been observed, indicating that Doxil is an active antitumor compound. Polyethyleneglycol-coated liposomes show a distinct advantage over previous liposome formulations directed at the RES and appear to be a promising drug delivery system for doxorubicin.

The effects of chemotherapy on morphology, cellular proliferation, apoptosis and oncoprotein expression in primary breast carcinoma.

Abstract: The use of chemotherapy as a form of primary treatment for breast cancer is increasing and, as a result, more resection specimens contain tumours which have been exposed to cytotoxic drugs. We have studied the effects of chemotherapy on the tumour morphology and various biological features of breast carcinoma in a group of 35 patients. These were a group who responded to treatment in a clinical study of the use of primary chemotherapy designed to reduce tumour bulk prior to surgery. Characteristic morphological changes, temporally related to the administration of cytotoxic agents, are seen. The malignant cells become enlarged with vacuolated cytoplasm and vesicular nuclei containing prominent nuclei; occasionally the nuclei were angular and hyperchromatic. These features are interpreted as degenerative in nature. In 15 cases sufficient material was present in the pretreatment biopsies to compare the grade of the tumours before and after chemotherapy: changes were found in six tumours. Cytotoxic drugs do not induce a consistent pattern of change in the proliferation and apoptotic indices of individual tumours, but there is a tendency to reduce proliferative activity over all the tumours as a group. It was also found that chemotherapy is capable of modifying the expression of the oncoproteins c-erbB-2 and p53 in a minority of cases of breast cancer, usually resulting in an acquisition of immunoreactive oncoprotein. It is important to be aware of these effects when studying breast carcinomas removed after chemotherapy.

Adenosine triphosphate-dependent transport of doxorubicin, daunomycin, and vinblastine in human tissues by a mechanism distinct from the P-glycoprotein.

Abstract: Previous studies have demonstrated that a human glutathione conjugate transporter, designated as dinitrophenyl-S-glutathione ATPase (DNP-SG ATPase), catalyzed ATP hydrolysis in the presence of several amphiphilic compounds other than glutathione conjugates (Singhal, S. S., R. Sharma, S. Gupta, H. Ahmad, P. Zimniak, A. Radominska, R. Lester, and Y. C. Awasthi. 1991. FEBS [Fed. Eur. Biochem. Soc.] Lett. 281:255-257). We now demonstrate that DNP-SG ATPase purified from human lung and erythrocyte membranes catalyzed the hydrolysis of ATP in the presence of doxorubicin and its metabolites. Doxorubicin-stimulated ATP hydrolysis by DNP-SG ATPase was saturable with respect to doxorubicin (Km 1.2 and 2.8 microM for the lung and erythrocyte enzymes, respectively). Antibodies against DNP-SG ATPase immunoprecipitated the ATP hydrolyzing activity stimulated by doxorubicin, its metabolites, and glutathione conjugates. Inside our vesicles prepared from erythrocyte membranes took up doxorubicin, daunomycin, and vinblastine in an ATP-dependent manner. The uptake was linear with respect to time and vesicle protein, was dependent on ATP and magnesium, was inhibited by heavy metal salts or by heating the vesicles, and was sensitive to both osmolarity and orientation of the vesicles. The transport had an activation energy of 13 kcal/mol, was saturable with respect to both doxorubicin and ATP (Km values of 1.8 microM and 1.9 mM, respectively), and was competitively inhibited by glutathione conjugates as well as by a number of amphiphiles such as daunomycin or vinblastine. Transport was diminished upon coating the vesicles with antibodies against DNP-SG ATPase. Incorporation of increasing amounts of purified DNP-SG ATPase into the vesicles resulted in a linear increase in transport of doxorubicin. These studies demonstrated for the first time that a membrane protein that catalyzed the transport of anionic amphiphilic molecules such as glutathione conjugates could also mediate the transport of weakly cationic antitumor antibiotic, doxorubicin. Notably, the Km of transport was in the range of doxorubicin concentration achievable in human serum after intravenous dosing of doxorubicin.

Organ-Specific Modulation of Steady-State mdr Gene Expression and Drug Resistance in Murine Colon Cancer Cells

Abstract: BACKGROUND The major cause of death from cancer is metastases that are resistant to conventional therapies. The resistance of metastatic tumor cells to chemotherapy can be caused by their intrinsic properties, such as increased expression of the mdr genes. PURPOSE The purpose of our present study was to determine some of the mechanisms by which the organ microenvironment influences the response of tumor cells to chemotherapy. METHODS Murine CT-26 colon cancer cells growing in continuous culture (parental cells) were harvested and injected subcutaneously into the lateral flank (to produce subcutaneous tumors) or the lateral tail vein (to produce experimental lung metastases) of 10 8-week-old syngeneic male BALB/c mice. Seven days after tumor-cell injection, the mice were given intravenous injections of either doxorubicin (10 mg/kg) or 0.9% NaCl (controls). This in vivo injection was repeated 7 days later. Mice with subcutaneous tumors and lung metastases were killed by cervical dislocation on day 21, and tumor samples from control mice were harvested and adapted to culture. The sensitivity of the cultured cells to doxorubicin and fluorouracil (5-FU) was determined at multiple time points. Levels of mdr-1 DNA were measured by slot-blot and Southern-blot analyses. mdr mRNA expression levels were measured by Northern-blot analysis using mdr-1- and mdr-3-specific hybridization probes, and P-glycoprotein level was determined by fluorescence-activated cell sorting using different monoclonal antibodies. RESULTS Treatment with doxorubicin produced 80% growth inhibition of CT-26 subcutaneous tumors but had little effect on the number (and size) of experimental lung metastases. Collectively, the results suggest that the multidrug-resistant phenotype developed in CT-26 cells growing in the lung environment. Cultures established from lung metastases were initially resistant to doxorubicin (but not to 5-FU) and showed elevated expression of mdr-1 mRNA transcripts and P-glycoprotein. This resistance could be overcome by verapamil and disappeared after 21 days in culture. No mdr gene amplification was detected. The expression level of mdr-specific mRNA (predominance of mdr-1) and P-glycoprotein was directly associated with resistance to doxorubicin. CONCLUSIONS Results of this study have demonstrated that the in vivo sensitivity of murine CT-26 colon carcinoma cells to doxorubicin depends on the organ environment. The organ environment can influence the P-glycoprotein-mediated multidrug-resistant phenotype in tumor cells, and the increased expression of P-glycoprotein is transient; once removed from the environment (lung), the cell's resistance reverts to that of the sensitive parent cells.

MDR1 Gene Expression: Its Effect on Drug Resistance to Doxorubicin in Human Hepatocellular Carcinoma Cell Lines

Abstract: Background Hepatic tumors are resistant to many chemotherapeutic agents. Although elevated MDR1 (also known as PGY1) gene expression has been shown in such tumors, no direct association has been established between the gene expression and multidrug resistance. Purpose To evaluate the role of the MDR1 gene in the drug resistance of hepatoma, we tested nine human hepatoma cell lines for their expression of the MDR1 gene. Methods We measured the MDR1 messenger RNA (mRNA) expression by RNA slot-blot analysis and by immunocytochemical staining with a P-glycoprotein-specific monoclonal antibody, MRK16. The in vitro chemosensitivity of these cell lines to fluorouracil, doxorubicin, mitomycin C, cisplatin, and etoposide (VP-16) was determined using the MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide) colorimetric assay. For doxorubicin cytotoxicity, we also tested the potentiating effect of several multidrug resistance-reversing agents. Results Slot-blot analysis and immunocytochemistry showed that two cell lines expressed high levels of MDR1 mRNA, one expressed an intermediate level, and all others were low expressors. The MTT assay results showed that all cell lines tested were generally resistant to chemotherapeutic agents. The assay area under the curve (AUC) was within a clinically achievable range only for VP-16 in one of nine cell lines. When the IC50 values were compared among the cell lines, the results revealed a close association with the MDR1 gene expression only for doxorubicin resistance. Verapamil and quinidine lowered the IC50 values of doxorubicin for MDR1-positive cell lines. The lowered assay AUC levels for both reversing agents, however, were still higher than the clinically achievable range. Conclusion These results indicate that the MDR1 gene probably has a role in doxorubicin resistance in hepatocellular carcinoma and that the resistance can be overcome by some multidrug resistance-reversing agents. Implications Some widely used anticancer agents might be ineffective for treating hepatocellular carcinoma in clinical situations even when combined with reversing agents.

Comparison of 5-fluorouracil, doxorubicin and mitomycin C with 5-fluorouracil alone in the treatment of pancreatic-biliary carcinomas.

Abstract: In this multicenter randomized trial, the efficacy of combination chemotherapy using 5-fluorouracil, doxorubicin and mitomycin C (arm A) was compared with that of 5-fluorouracil alone (arm B) in 81 patients with nonresectable carcinomas of the pancreas or biliary tract. There were no significant differences between treatment arms regarding the median time to progressive disease, median survival time, palliative effects or toxicities. It was concluded that combination chemotherapy is feasible but cannot be recommended.

Liposomal doxorubicin in the treatment of advanced AIDS-related Kaposi sarcoma

Abstract: Neither single-agent therapy nor any combination treatment has been satisfactory enough to be regarded as standard in systemic advanced Kaposi sarcoma. In an attempt to achieve high efficacy in combination with low toxicity, we used a new liposomal formulation of doxorubicin. Pharmacologic data had established a long plasma half-life, an increased accumulation in tumor tissue, and a decrease in uptake by tissues such as liver, spleen, and bone marrow. In a phase I/II open-label, dose-escalating trial 40 male AIDS patients with advanced Kaposi sarcoma were enrolled to receive intravenous "stealth" liposomal doxorubicin biweekly at doses of 10 mg/m2 (n = 10), 20 mg/m2 (n = 27), and 40 mg/m2 (n = 3). The median CD4 count at baseline was 25/microL. After six cycles (12 weeks), 39 patients were evaluable. Three patients (7.5%) showed a complete response, which was histologically confirmed. A partial response was documented in 33 patients (85%). Stable disease was observed in three patients (7.5%). During a median treatment duration of 25 weeks, four patients developed stomatitis (10%), and four patients (10%) experienced alopecia. The most frequent hematologic toxicity was neutropenia. Grade 4 neutropenia was seen in 42.5%, and grade 3 toxicity was seen in 30%. Toxicity was dose-dependent and more frequent in the 40 mg/m2 stratum. During a median observation period of 25 weeks, opportunistic infections occurred in 57.5% of the patient population. We conclude that liposomal doxorubicin at dose levels of 10 and 20 mg/m2 is safe and effective for treatment of advanced Kaposi sarcoma in AIDS. A controlled trial comparing liposomal doxorubicin to conventional combination therapy is underway.

Preoperative and postoperative adjuvant combination chemotherapy for adults with high grade soft tissue sarcoma.

Abstract: BACKGROUND Patients with high grade soft tissue sarcoma greater than or equal to 10 cm have a 3-year disease-free survival of approximately 30%. There is no convincing evidence, however, that postoperative adjuvant chemotherapy is beneficial. Preoperative chemotherapy has theoretical advantages over postoperative chemotherapy. METHODS Twenty-nine evaluable patients with primary or recurrent high grade, nonmetastatic, soft tissue sarcoma were treated with two preoperative cycles of cyclophosphamide 500 mg/m2, doxorubicin 60 mg/m2, and DTIC 1000 mg/m2 before definitive surgery and radiation. Clinical and radiologic assessment of response to chemotherapy was performed preoperatively, and the resected specimen was examined for treatment effects. Patients who did not progress during preoperative therapy were eligible to receive four additional cycles of chemotherapy. Disease-free and overall survival rates of study patients were compared with two cohorts of historic controls. RESULTS Although subjective changes in the firmness of some tumors were observed, only one patient met the criteria for partial response (3%, 2-sided 95% confidence interval = < 1-17%). Intratumoral hemorrhage, cystic necrosis, and liquefaction were observed regularly, and three tumors were more than 90% necrotic. Toxicity of the chemotherapy was acceptable, but patients were reluctant to receive postoperative therapy. The median time free from distant metastasis was 28 months; median survival was 35 months. These results were not superior to the experience with no chemotherapy, or with postoperative doxorubicin. CONCLUSIONS Adjuvant chemotherapy for patients with soft tissue sarcoma remains investigational. There is a strong rationale, however, for continued investigation of preoperative chemotherapy for high risk patients using doxorubicin and ifosfamide with colony stimulating factor support. Development of sensitive and specific methods to assess response to preoperative chemotherapy is needed.

Effects of hyaluronidase on doxorubicin penetration into squamous carcinoma multicellular tumor spheroids and its cell lethality

Abstract: Doxorubicin is an anticancer agent widely used in the treatment of human cancer. The major limitation of this drug governing the cell-killing effect appears to be its poor penetration into a tumor mass. We have studied the effects of hyaluronidase on the penetration and cell-killing effect of doxorubicin using multicellular tumor spheroids (MTS). MTS approximately 500 μm in diameter were produced by a liquid-overlay culture technique from PC-10 lung and HEp-2 laryngeal squamous carcinoma cell lines. Cells in MTS and monolayer were exposed to hyaluronidase for various lengths of time; this was followed by a 1-h resting interval and a subsequent 1-h exposure to doxorubicin. MTS and monolayer cells were then trypsinized to a single-cell suspension and subjected to clonogenic assay. Hyaluronidase at a concentration of 25 U/ml or 250 U/ml was nontoxic to the monolayer cells. For PC-10 MTS, pretreatment with 25 U/ml hyaluronidase for 24 h and 72 h resulted in approximately 20% increases in Doxorubicin cell killing at the median (IC50) dose as compared to doxorubicin alone. HEp-2 MTS were more sensitive to the hyaluronidase pretreatment. Thus, a 1-h exposure to the enzyme produced a 40% increase in doxorubicin-induced cell death at the IC50 dose. A fluorescence microscopic study revealed that a 1-h exposure of MTS to doxorubicin produced doxorubicin fluorescence only in the one or two outer layers of MTS. When MTS were pretreated with hyaluronidase, there was enhanced penetration of doxorubicin fluorescence into the MTS core. Hyaluronidase-induced enhancement of Doxorubicin penetration and its cell-killing effect is dependent on the exposure time and tumor cell origin. These data suggest that anecdotal reports of hyaluronidase-enhanced activity of preclinical chemotherapy deserve a controlled trial.

Linalool, a plant-derived monoterpene alcohol, reverses doxorubicin resistance in human breast adenocarcinoma cells.

Abstract: Essential oils from various aromatic plants have been reported to exert chemopreventive and/or antitumor effects. In addition, a number of studies have shown the ability of chemopreventive phytochemicals to increase the sensitivity of cancer cells to conventional anticancer drugs. The success of chemotherapeutic agents is often hindered by the development of drug resistance, with multidrug resistant (MDR) phenotypes reported in a number of tumors, generally involving reduced intracellular drug accumulation due to increased drug efflux by membrane transporters. In the present study, the effects of linalool (LIN), a monoterpene alcohol found in the essential oils from many aromatic plants, on the growth of two human breast adenocarcinoma cell lines, MCF7 WT and multidrug resistant MCF7 AdrR, were investigated, both as a single agent and in combination with doxorubicin (DOX). The results reported here show that LIN only moderately inhibits cell proliferation; interestingly, however, subtoxic concentrations of LIN potentiate DOX-induced cytotoxicity and pro-apoptotic effects in both cell lines. A significant synergism can be observed in MCF7 AdrR cells, which may be due, at least in part, to the ability of LIN to increase DOX accumulation and to induce a decrease in Bcl-xL levels. In summary, the results of the present study suggest that LIN may improve the therapeutic index of anthracyclines in the management of breast cancer, especially in MDR tumors.

MDR-1 expression and response to vincristine, doxorubicin, and dexamethasone chemotherapy in multiple myeloma refractory to alkylating agents.

Abstract: PURPOSETo assess whether the presence of enhanced multiple drug resistance (MDR)-1 gene expression in multiple myeloma (MM) patients predicts survival, as well as response to vincristine, doxorubicin, and dexamethasone (VAD) chemotherapy.PATIENTS AND METHODSSixty-three MM patients refractory to alkylating therapy were studied. The presence of the MDR-1 gene product, a 170-kd glycoprotein (P-170), was analyzed in bone marrow plasma cells by means of the alkaline phosphatase (APAAP) technique using the P-170-specific monoclonal antibody (MoAb) C219. The prognostic value of MDR-1 gene expression, examined before VAD treatment, was compared with other established prognostic factors including beta 2-microglobulin, albumin, lactate dehydrogenase (LDH), and the plasma cell labeling index.RESULTSFifty-nine percent of all samples were P-170-positive. No association could be demonstrated between response to VAD and MDR-1 gene expression (chi 2 P = .359), in contrast to high serum beta 2-microglobulin levels, which ...

Hyperthermia induces doxorubicin release from long-circulating liposomes and enhances their anti-tumor efficacy

Abstract: Purpose: To examine the possibility that hyperthermia would accelerate drug release from long-circulating liposomes, and enhance their antitumor activity. Methods and Materials: Liposomes were prepared by thin film hydration technique. Hyperthermia was induced by ultrasound apparatus and a water bath heating system. The antitumor efficacy of treatment against RIF-1 tumor in C3H mice was evaluated by the tumor growth delay assay. Results: In vitro drug release experiments demonstrated that increase in temperature from 37°C to 41°C resulted in about a sixfold increase in doxorubicin (DOX) release in a 1-h period. Increasing the temperature to 43°C, resulted in only a modest additional drug release. Drug uptake studies showed that local hyperthermic treatment immediately following the drug administration dramatically enhanced Stealth liposome-encapsulated doxorubicin (S-DOX) uptake by tumors, but did not do so for free DOX. At 42°C and at a dose of 10 mg/kg, the accumulation of S-DOX was about 10-fold and 2.5-fold higher than that with free drug and S-DOX at 37°C, respectively. The antitumor efficacy study confirmed our hypothesis that the addition of hyperthermia to the treatment of RIF-I tumors with doxorubicin encapsulated in long-circulating liposomes would enhance antitumor effects. Two hyperthermia treatments given at 24-h intervals appeared to be the most promising method of combining heat and long-circulating liposomes. The increased antitumor activity was not accompanied by increased toxicity, as determined by the body weight of the mice. Conclusion: Local hyperthermic treatment is able to accelerate DOX release from long-circulating liposomes, increase tumor uptake, and enhance their antitumor efficacy. The combination of local hyperthermia and long-circulating liposomes appears to show considerable promise in the treatment of localized diseases.

Protection against doxorubicin-induced alopecia in rats by liposome-entrapped monoclonal antibodies.

Abstract: Alopecia is a common side effect of several anti-cancer drugs, including doxorubicin. Based on our recent observation that a monoclonal antibody (MAD11) directed against this anthracycline inhibits the systemic toxic effect of the drug in mice, we investigated the possibility that MAD11 administered topically might protect against doxorubicin-induced alopecia. In 31 of 45 young rats treated intraperitoneally with doxorubicin, alopecia was completely prevented by topical treatment of the skin with liposome-incorporated anti-doxorubicin monoclonal antibody. This type of treatment might find relevance in preventing anthracycline-induced alopecia in cancer patients. Our findings also provide the first demonstration that liposome-entrapped monoclonal antibodies are capable of penetrating the stratum corneum of the skin without losing their function.

Antitumor Activity of Free and Liposome-entrapped Annamycin, a Lipophilic Anthracycline Antibiotic with Non-Cross-Resistance Properties

Abstract: The lipophilic anthracycline antibiotic annamycin (Ann) was entrapped in liposomes of different size [median diameter: 1.64 microns, multilamellar liposomal Ann (L-Ann); 0.030 micron, small unilamellar Ann (S-Ann)] with > 90% entrapment efficiency and tested in vitro against four pairs of sensitive and multidrug-resistant (MDR) tumor cell lines and in vivo by the i.v. route in five tumor models: advanced s.c. B16 melanoma; s.c. M5076 reticulosarcoma; lung metastases of Lewis lung carcinoma; and s.c. KB and KB-V1 xenografts in nude mice. Predetermined optimal doses of the different formulations were used and the results were compared with doxorubicin (Dox). In vitro, Ann, either in suspension in 10% dimethyl sulfoxide (F-Ann) (1 mg/ml) or entrapped in liposomes, was able to partially overcome resistance in all four pairs of sensitive and MDR KB, 8226, P388, and CEM cell lines (resistance indexes 63, 269, 333, and 356 for Dox versus 4, 5, 19, and 8.7 for L-Ann, respectively). In vivo, both F-Ann and liposome-entrapped Ann were slightly more effective than Dox in inhibiting the growth of advanced s.c. B16 melanoma tumors. L-Ann was markedly more effective than Dox and moderately more effective than F-Ann in prolonging the life span of animals bearing s.c. M5076 and lung metastases of Lewis lung carcinoma tumors. All drugs were equally effective at optimal doses in delaying the growth of s.c. KB xenografts, whereas all Ann formulations were markedly more effective than Dox in delaying the growth of s.c. KB-V1 (MDR) xenografts. In all in vivo experiments, S-Ann was consistently more effective than L-Ann and L-Ann was more effective than F-Ann. These results indicate that (a) Ann is more effective than Dox by the i.v. route against several tumor models and that MDR tumors are partially not cross-resistant to Ann both in vitro and in vivo, (b) liposomes enhance the in vivo antitumor properties of Ann, and (c) small liposomes are more effective than large liposomes in enhancing Ann antitumor activity.

Effect of vitamin A administration on resistance of rat heart against doxorubicin-induced cardiotoxicity and lethality.

Abstract: The peroxyl radical-scavenging activity of vitamin A has been exploited to obtain protection against peroxidative damages induced in rat heart by administration of an acute dose of doxorubicin (10 mg/kg, in vein). Peroxidative lesions were evaluated by both biochemical and histological assays, 48 hr after the injection of doxorubicin. Heart tissue from rats receiving doxorubicin showed a marked increase of protein carbonyl levels, and of membrane conjugated dienes, as well as a decrease of membrane protein thiols. Abnormal chemistries, including a large increase of the activity of serum lactate dehydrogenase and creatine phosphokinase, an index of the myocardial damage caused by doxorubicin, were also observed. Pretreatment of rats with 25 I.U./kg b.wt. of vitamin A, once a day for 2 days, before injecting doxorubicin, substantially reduced the peroxidative damage to heart lipids and proteins, and markedly lowered the serum values of lactate dehydrogenase and creatine phosphokinase to values close to those of control rats. The significant prevention of doxorubicin-induced cardiomyopathy by vitamin A was evident from the histopathological pattern observed after light microscopy. The dosage of vitamin A useful to obtain the protective effect appears safe and does not injure the liver, as indicated by light microscopy of the tissue. A survival study, carried out by injecting rats with a single injection of 10 mg/kg of doxorubicin, showed that pretreatment with 25 I.U. of vitamin A per kg significantly increased survival rate of the animals.(ABSTRACT TRUNCATED AT 250 WORDS)