Liposome-encapsulated drug delivery is a methodology that has been evolving over the past 30 years. A number of liposome-encapsulated anthracycline products are in development and two, pegylated liposomal doxorubicin
ABSTRACT: Liposome-encapsulated drug delivery is a methodology that has been evolving over the past 30 years. A number of liposome-encapsulated anthracycline products are in development and two, pegylated liposomal doxorubicin (PEG-LD) (Doxil) and liposomal daunorubicin (Dauno-Xome), are approved for the treatment of AIDS-related Kaposis sarcoma. Preclinical studies show PEG-LD to be at least as effective as traditional or free doxorubicin (Adriamycin) in a variety of tumor models. Pharmacokinetic studies reveal differences between PEG-LD and doxorubicin, with PEG-LD having a higher area under the concentration-time curve (AUC), lower clearance rate, and smaller volume of distribution. In addition, PEG-LD was found to selectively accumulate in cutaneous Kaposis sarcoma lesions of AIDS-infected individuals when compared with adjacent normal skin. Accumulating data have led to a proposed mechanism of PEG-LD accumulation in tumors: long-term circulating liposomes pass through gaps/defects in newly formed blood vessels and enter the tumor interstitium. Liposome breakdown within tumors releases doxorubicin molecules that travel deeper into the tumor, bind to nucleic acids, and result in tumor-cell killing. The ability of PEG-LD liposomes to remain intact while in circulation, retaining most of the doxorubicin in encapsulated formulation, is believed to be responsible for the reduced toxicity seen with this agent without sacrificing efficacy. [ONCOLOGY 11(Suppl 11):11-20, 1997]
Soon after Bangham and Horne first described liposomes in the mid- 1960s as closed vesicular structures able to envelop water-soluble molecules, pharmacologists recognized their potential value in drug delivery. The rationale was simpleuse liposomes as a safe vehicle for delivering drugs more specifically to sites of disease while limiting exposure of normal tissues. The envelopment of cytotoxic antitumor agents was of particular interest because these drugs generally have a narrow therapeutic window, ie, dose-limiting side effects limit their therapeutic utility.
Among the dozens of liposome-encapsulated antitumor agents studied in animal models, the anthracycline antibiotics, in particular, doxorubicin and daunorubicin, emerged as benefiting substantially from liposome encapsulation. Animals were able to tolerate greater doses of a variety of formulations of liposome-encapsulated doxorubicin (Doxil) with antitumor activity, in general, being maintained. Although this safety advantage was considered meaningful, attempts to improve antitumor activity by actually targeting encapsulated drugs to tumors were frustrated by rapid removal of the liposomes from blood by elements of the mononuclear phagocyte system, fixed macrophages residing in liver, spleen, lung, and bone marrow. It is believed that the binding of plasma proteins (lipoproteins, immunoglobulins, complement) to the liposome surface triggers this rapid macrophage uptake.
Despite the lack of true targeting, internalization of liposome-encapsulated anthracyclines by mononuclear phagocyte system cells was found to diminish exposure of certain tissues to the toxic effects of such drugs. For example, doxorubicin-related nausea/vomiting and cardiomyopathy are believed to be related to the drugs peak levels in plasma. In theory, liposome encapsulation results in sequestration of the majority of an injected dose in the mononuclear phagocyte system, thus attenuating the initial plasma levels of free drug and improving safety. The drug is eventually released from mononuclear phagocyte system cells and is distributed to peripheral tissues in free (ie, unencapsulated) form. If this theory is true, the pharmacokinetic pattern would mimic that of doxorubicin administered as a prolonged infusion, a regimen known to reduce drug-related side effects. Indeed, it has been shown that administration of liposome-encapsulated doxorubicin reduces the drugs acute and chronic toxicities in preclinical animal models. Moreover, results from animal models indicate that doxorubicin delivered in this fashion retains its activity against systemic tumors.
The pharmacokinetics and safety of various clinical formulations of liposomal doxorubicin have been reported in the scientific literature.[2,6-24] Clinical pharmacokinetic measurements confirm that conventional liposome formulations are cleared rapidly from plasma. These data also suggest that a considerable amount of encapsulated doxorubicin is released into plasma prior to mononuclear phagocyte system uptake.[10,21]
Armed with the knowledge that mononuclear phagocyte system uptake can provide favorable safety advantages for encapsulated doxorubicin, formulation scientists began to optimize liposome carriers for this purpose. As shown in Figure 1, the first major branch of the liposome anthracycline family tree was represented by these Mononuclear Phagocyte System Targeted formulations. Two alternative formulation approaches (subbranches) soon emerged.
The first approach relied upon acidic lipids incorporated into the liposome bilayer, such as cardiolipin (CL) and egg phosphatidyl glycerol (EPG), to bind doxorubicin, which is positively charged at physiological pH, to the membrane itself.[8,12] Formation of such ion-pairs between the drug and an acidic membrane component provided a stable association resulting in robust formulations that were stable in vitro and could be freeze-dried for long-term storage.
The second approach, represented by TLC D-99, used true encapsulation of doxorubicin into the aqueous compartment of the liposome and employed a clever technique to circumvent the problem of leakage. In this case doxorubicin is loaded into the liposomes (in a hospital pharmacy) immediately prior to administration by adding an aqueous solution of doxorubicin (at neutral pH 7.0) to liposomes containing a low pH internal buffer (pH 4.0). The pH gradient thus formed across the liposome membrane leads to movement of doxorubicin into the liposomes. Once inside, the low pH environment traps the drug, preventing it from leaking out as long as the pH gradient is maintained.
The ion-pair formulations have been tested clinically but have not progressed beyond phase I/II studies. TLC-D99 is in advanced phase III trials in metastatic breast cancer.
Recognizing that mononuclear phagocyte system uptake represented the main obstacle to targeting, another branch of liposome scientists attempted to formulate liposomes that could resist binding/interaction with plasma proteins (opsonization), thus prolonging their blood residence times and targeting potential. Early work suggested that a modest degree of mononuclear phagocyte system avoiding activity could be obtained by formulations composed of high phase transition lipids and cholesterol. Size was also a critical parameter: the smaller the liposome, the longer it circulated. This pure lipid subbranch arrived at two formulations of small diameter (approximately 50 nm), one composed of DSPC/cholesterol and the other of sphingomyelin/cholesterol, both of which showed relatively slow mononuclear phagocyte system clearance. DaunoXome, a DSPC/cholesterol formulation of daunorubicin, is the only product to emerge from this pure lipid approach. DaunoXome is approved in the US and Europe for the treatment of AIDS-related Kaposis sarcoma.
Not satisfied with the plasma longevity achieved by the pure lipid formulations, another branch of liposome scientists explored the possibility of coating the surface of liposomes with inert materials designed to camouflage the liposome from the bodys host defense systems. The biological paradigm for this surface modified subbranch was the erythrocyte, a cell which is coated with a dense layer of carbohydrate groups, and which manages to evade immune system detection and to circulate for several months before being removed by the same type of cell responsible for removing liposomes. The first breakthrough came in 1987 when a glycolipid (the brain tissue-derived ganglioside GM1) was identified which, when incorporated within the lipid matrix, allowed liposomes to circulate for many hours in the blood stream. A second glycolipid, phosphatidylinositol, was also found to impart long plasma residence times to liposomes and, because it was extracted from soy beans not brain tissue, was believed to be a more pharmaceutically acceptable excipient.
A major advance in the surface modified subbranch was the development of polymer-coated liposomes. Polyethylene glycol modification had been used for many years to prolong the half-lives of biological proteins (such as enzymes and growth factors) and to reduce their immunogenicity. Several groups of liposome scientists reported in the early 1990s that pegylated-coated liposomes circulated for remarkably long times after intravenous administration. Half-lives on the order of 24 hours were seen in mice and rats and over 30 hours in dogs. The term stealth was applied to these liposomes because of their ability to evade interception by the immune system in much the same way as the stealth bomber was able to evade radar. Pegylated liposomal doxorubicin (PEG-LD) (Doxil) is the first product to emerge from the surface modified liposome subbranch. It too is approved in the US and Europe for treatment of Kaposis sarcoma.
The efficacy of PEG-LD has been evaluated in a variety of different tumor models, including several human xenograft models.[30-36] In every model examined, PEG-LD was more effective than the same dose of free doxorubicin in inhibiting or halting tumor growth, in effecting cures and/or in prolonging survival times of the tumor-bearing animals. In most cases, all three endpoints were improved by PEG-LD, and in no case was PEG-LD less effective than doxorubicin. Not only was PEG-LD more active in both solid and dispersed tumors, it was also found to be more effective than doxorubicin in preventing spontaneous metastases from two different intramammary implanted tumors in mice. These findings also are supported by studies performed with PEG-LD in several murine tumor models and a human xenograft model (Figure 2).
The efficacy of free doxorubicin in these animal models was generally limited by its toxicity at high doses. Therefore, the ability to use PEG-LD at higher doses offers a therapeutic advantage. In addition, pharmacokinetic and tissue distribution studies suggest that the greater persistence, particularly in tumor tissue, achieved with PEG-LD compared with conventional doxorubicin also contributes a therapeutic advantage.
Furthermore, PEG-LD was found to be significantly more effective than conventional (non-Stealth) liposomal doxorubicin, demonstrating the impact of the long-circulating Stealth liposome. Based on the results of these nonclinical studies, PEG-LD appears to be an effective agent for the treatment of both solid and dispersed tumors.
Population pharmacokinetic analysis has been conducted on a group of 83 patients receiving PEG-LD at doses ranging from 10 to 60 mg/m2 (17 females, 66 males).[37,38] Between 10 and 40 mg/m2, PEG-LD pharmacokinetics were linear; at dosages above 40 mg/m2, PEG-LD displayed nonlinear pharmacokinetics as evidenced by a disproportionate increase in the area under the plasma concentration-time curve (AUC) with increasing dose amounts. In general, drugs that display nonlinear pharmacokinetics have a potential to accumulate to toxic levels in the plasma if not monitored regularly (eg, phenytoin). This is not a concern in the case of PEG-LD, because the drug is administered at a minimum of once every 3 weeks, after which time no drug is detectable in the plasma of patients. Table 1 lists selected pharmacokinetic parameters for all 83 patients. No evidence of drug accumulation is seen at dose intervals of greater than or equal to 3 weeks. Utilizing the fitted pharmacokinetic parameter results from this analysis, simulated plasma concentration versus time profiles of PEG-LD were generated at doses of 10 to 60 mg/m2 (Figure 3). The nonlinearity of PEG-LD pharmacokinetics at higher doses is most evident at doses greater than 40 mg/m2.
No correlations were observed between pharmacokinetic parameters and either age, weight, body surface area, tumor type, sex, renal (as determined by serum creatinine) or hepatic function (as determined by total bilirubin levels).
Amount of Nonliposomal Doxorubicin in Plasma
Several lines of evidence support the conclusion that after IV administration of PEG-LD, the majority of the doxorubicin in plasma (between 93% and 99%) is encapsulated within the liposome. The most convincing data come from work by Gabizon et al who conducted a pilot pharmacokinetic trial of PEG-LD. In this study, the fraction of both the liposome-encapsulated and the free, nonliposomal drug in circulation after PEG-LD administration were quantitated directly using a Dowex column separation method that is able to accurately and reproducibly quantitate greater than or equal to 7% free drug in plasma. Using this method, it was determined that essentially all the doxorubicin measured in plasma was liposome-associated (Figure 4). These findings suggest that at least 90% to 95% of the doxorubicin measured in plasma, and possibly more, is liposome-encapsulated.
The amount of doxorubicin that remains liposome-associated while circulating in plasma is an important factor that deserves further emphasis from a safety perspective. Acute adverse reactions associated with doxorubicin administration, including nausea and vomiting and chronic cardiotoxicity, are believed to be directly related to peak concentrations of the drug in plasma. As pointed out above, PEG-LD remains intact while in the circulation retaining virtually all of the doxorubicin in encapsulated form. Although total plasma levels of doxorubicin may be relatively high for several days after PEG-LD administration, the majority of the dose is sequestered within the liposome during this period and, thus, is not bioavailable to distribute (as free drug molecules) to tissues, including the gastrointestinal tract and the myocardium. With respect to the level of available drug in plasma, PEG-LD administration more closely resembles that of a 96-hour continuous infusion of doxorubicin than the usual 30-minute infusion. Prolonging the infusion of doxorubicin is known to reduce cardiotoxicity and gastrointestinal irritation.
Comparison of Pharmacokinetic Parameters: PEG-LD vs Doxorubicin
According to literature reports, an IV bolus injection of doxorubicin in humans produces high plasma concentrations of doxorubicin that decline quickly due to rapid and extensive distribution into tissues. Apparent volumes of distribution range from 1,400 to 3,000 L, reflective of the drugs extensive tissue distribution. The doxorubicin plasma concentration-time curve in humans is biphasic, with a distribution half-life of 5 to 10 minutes and a terminal phase elimination half-life of 30 hours.[42-44] A triphasic curve has also been described with a terminal plasma half-life of approximately 30 hours. Clearance of doxorubicin after its administration ranges from 24 to 73 L/hour. No accumulation in plasma occurs after repeated injections.[42-44]
The pharmacokinetics of PEG-LD are significantly different from those reported for doxorubicin. Administration of PEG-LD results in a significantly higher doxorubicin AUC, lower rate of clearance (approximately 0.1 L/hour) and smaller volume of distribution (5 to 7 L) relative to administration of doxorubicin. The first phase of the biexponential plasma concentration-time curve after PEG-LD administration is relatively short (approximately 5 hours), and the second phase, which represents the majority of the AUC, is prolonged (half-life, 50 to 55 hours).
Cmax after administration of PEG-LD is 15-fold to 40-fold higher than after the same dose of doxorubicin, and the ratio quickly increases as doxorubicin is rapidly cleared from circulation. Importantly, the vast majority of the total plasma doxorubicin remains liposome-encapsulated after PEG-LD treatment. Because of the high percentage of liposome encapsulation in PEG-LD, the amount of free (ie, bioavailable) drug in the plasma appears to be significantly lower than that measured after administration of an equal dose of doxorubicin.
This conclusion is supported by calculations which derive the apparent concentration of free doxorubicin from the reported relationship between doxorubicinol and doxorubicin concentrations in plasma. For example, 5 minutes after the end of the infusion of a 20 mg/m2 dose of PEG-LD, the mean doxorubicinol level was approximately 22 ng/mL. Using the doxorubicinol:doxorubicin concentration ratio reported in the literature, predicted free doxorubicin concentration at this time point would be 54 ng/mL in PEG-LD-treated patients (the total plasma concentration measured at this time point was 8,863 ng/mL). Comparatively, patients in the Northfelt et al study who received a 20 mg/m2 dose of doxorubicin had initial plasma concentrations of doxorubicin of approximately 500 ng/mL.
Doxorubicin Levels in Kaposis Sarcoma Lesions
Biopsies of Kaposis sarcoma (KS) lesion tissue and adjacent normal skin were obtained in 22 patients (Table 2). Doxorubicin levels in KS lesions were higher than in normal skin in 20 of the 22 patients; in 14 patients normal skin levels were below the lower limit of quantitation (0.4 µg/g tissue), whereas all KS lesion levels were quantifiable. Forty-eight hours after PEG-LD administration, median doxorubicin levels in biopsies of KS lesions ranged from 3-fold to 16-fold higher than in normal skin from the same patients. The median doxorubicin concentration in KS lesions was 1.3 µg/g tissue in seven patients receiving 10 mg/m2 PEG-LD and 15.2 mg/g tissue in seven patients receiving 20 mg/m2 PEG-LD; normal skin concentrations were 0.4 µg/g tissue in the 10 mg/m2 dose group and 0.9 µg/g tissue in the 20 mg/m2 dose group. Biopsy data 48 hours after PEG-LD injection in the seven patients receiving 20 mg/m2 are shown in Figure 5. Ninety-six hours after drug treatment, KS lesion doxorubicin levels were three-fold and five-fold greater than in normal skin from the same patients in the 10 and 20 mg/m2 groups, respectively. At 96 hours, median doxorubicin concentrations in KS lesions were 4.3 and 3.3 µg/g tissue in the four patients receiving 10 mg/m2 and the four patients receiving 20 mg/m2 dose, respectively; median concentration in the normal skin was 1.4 µg/g tissue in the 10 mg/m2 dose group, and 0.7 µg/g tissue in the 20 mg/m2 group.
Although too few time points were studied to allow determination of an AUC for doxorubicin in KS lesions or normal skin, these data suggest that doxorubicin accumulates in KS lesions after PEG-LD treatment.
An understanding of the mechanisms by which liposome-encapsulated doxorubicin accumulates within solid tumors after PEG-LD administration, and how this deposition pattern and subsequent slow release of drug improve the antitumor activity of PEG-LD relative to treatment with the free drug, is now emerging (Figure 6).
Plasma stability and long plasma residence times are critical requirements: PEG-LD liposomes are intended to carry their payload of doxorubicin directly to tumors. So, any premature release of the drug, while the liposomes are still in route (ie, in the circulation), would detract from the total amount of encapsulated doxorubicin able to reach the desired target. This requirement highlights the importance of engineering plasma stability into PEG-LD liposomes. As mentioned earlier, conventional liposome formulations of doxorubicin have been shown to release a significant proportion of their payload into the bloodstream soon after injection.[10,21] Drug release appears to follow protein adsorption/intercalation into the liposome which disrupts the barrier properties of the membrane. Moreover, the liposomes, together with any remaining drug, are removed by cells of the mononuclear phagocyte system within several minutes to a few hours after injection. As a consequence of this rapid clearance, doxorubicin delivered in conventional liposomes has little opportunity to reach tumors in encapsulated form.
By virtue of the pegylated groups grafted to their surface, pegylated doxorubicin liposomes are stable in plasma and release very little drug while in the circulation (see discussion above). Moreover, the pegylated coating provides slow clearance; after a single injection, PEG-LD can be detected in the circulation for 2 to 3 weeks. Slow clearance kinetics provide an opportunity for these liposomes to reach sites of disease such as tumors. Measurements made in tumor-bearing animals and in cancer patients indicate that uptake of pegylated liposomes by tumors is also a slow process. In preclinical tumor models, the peak uptake of PEG-LD is reached 24 to 48 hours after injection.[32,47] In cancer patients given indium 111 encapsulated in pegylated liposomes of the same composition and size as PEG-LD, peak uptake in tumors is seen 48 to 72 hours after injection (Figure 7). (Stewart Simon, personal communication, May 20, 1997) Slow uptake in tumors highlights the importance of long circulations times; if liposomes are to have an opportunity to reach and enter tumors in significant numbers, they must circulate for periods of days after injection.
Liposomes extravasate through gaps in the endothelium of tumor vessels: Stealth liposomes of the same size and lipid composition as PEG-LD, but containing entrapped colloidal gold designed to serve as a marker to follow liposome distribution by microscopic techniques, have been shown to enter solid colon tumors implanted in mice and KS-like lesions in HIV-transgenic mice. In these mouse models, movement of liposomes from the vascular lumen into the tumor interstitium was visualized by light and electron microscopy. Transcytosis of liposomes from the lumen of blood vessels through endothelial cells and into the extravascular compartment of KS lesions was seen, as was intracellular uptake of liposomes by some spindle cells within lesions. However, these processes appear to be restricted to a minority of the particles entering the tumor. The vast majority of the liposomes were seen to enter through gaps in the endothelial cell wall. This finding is consistent with results reported by Yuan et al who used pegylated liposomes ranging in size from 100 to 600 nm to probe the cut-off size of the gaps present in a human adenocarcinoma xenograft implanted in nude mice. This tumor was permeable to liposomes up to 400 nm in diameter, suggesting that the cut-off size in this tumor is between 400 and 600 nm. Given their small size (85 nm) and long circulation times, pegylated doxorubicin liposomes would be expected to extravasate in tumors that exhibit gaps of such dimensions. Gaps/defects are known to be present in solid tumors[51,52] and in KS lesions.[53,54] Indeed, fluorescent pegylated liposomes of less than 100 nm in diameter have been visualized by video microscopy extravasating in real time into the interstitium of implanted tumors using window chamber models.[55-57]
Release of drug following extravasation: Encapsulated doxorubicin is released from the PEG-LD liposomes after extravasation in tumors. Several possible factors may contribute to liposome breakdown and drug release in tumors: 1) conditions present in the interstitial fluid surrounding tumors that may cause breakdown of the liposomes include low pH and lipases released from dead or dying tumor cells; 2) inflammatory cells (which are often found in tumors) may release factors that lead to liposome destabilization, such as enzymes or superoxide and other oxidizing agents; or 3) phagocytic cells residing in tumors which are known to engulf liposomes, may digest the lipid matrix intracellularly and release doxorubicin or its active metabolites back into the interstitial fluid. A combination of these possibilities may well be responsible for the observed release of doxorubicin after extravasation of PEG-LD liposomes in tumors.
The rate of release of doxorubicin within a tumor has yet to be measured directly. In order to do so, it would be necessary to separate encapsulated drug (ie, drug molecules that have not been released from intact liposomes) from free drug in a solid tissue. Although such a separation is possible in biological fluids (such as plasma), it is technically difficult to conduct in solid tissues such as tumors; the conditions needed for quantitative extraction of doxorubicin lead to liposome disruption. Despite the difficultly of directly measuring release kinetics, indirect methods suggest that the release of doxorubicin from PEG-LD liposomes occurs over a period of days to perhaps weeks following administration. In a recent study using a human pancreatic xenograft model in nude mice, Vaage et al showed that tumor levels of doxorubicin peak at 24 to 48 hours after PEG-LD, and fall slowly over a period of 1 week. These results suggest that the liposomes entering the tumor release their drug locally at quite a slow rate.
The improved antitumor activity of PEG-LD relative to a comparable dose of free doxorubicin can be partially attributed to these slow in situ release kinetics. After a dose of free doxorubicin, drug molecules enter the tumor as well as other tissues quickly, reaching maximal exposure (ie, peak concentration) within minutes. During the subsequent 24 hours, tumor doxorubicin concentration drops precipitously to undetectable levels. During this brief pulse of doxorubicin, those cells not exposed to a cytotoxic concentration for a sufficient amount of time, or which are not at a sensitive point in the cell cycle, can escape therapy and continue to proliferate.
A typical course of doxorubicin is given on a 3-week cycle. This length of time between injections is needed to allow for recovery from the hematologic toxicity associated with doxorubicin therapy. Following such a schedule, it is quite likely that tumor cells are exposed to cytotoxic levels of drug for only a few hours during the 3-week interval between injections. In the case of PEG-LD, which is also given on a 2 to 4 week cycle, not only does more drug reach the tumor, but, by virtue of the slow in situ release kinetics provided by the liposomes, tumor cells are exposed to drug over a period of several days to perhaps a week or more after a single dose. Such a release pattern may contribute to the antitumor response of PEG-LD.
Tumor cell penetration and cytotoxicity: Given its amphipathic nature, a doxorubicin molecule that is released from a liposome can quickly diffuse through surrounding fluids and connective tissue, enter tumor cells, bind to nucleic acids, and inhibit DNA synthesis. Indeed, it is quite likely that drug molecules released from PEG-LD can penetrate many cell layers into the tumor, well beyond the point that the liposome itself has reached. Early findings suggest that penetration of free drug in this fashion may be essential for antitumor activity of PEG-LD.
As mentioned above, microscopic observations indicate that liposomes extravasate in tumors at particular sites, primarily through vessels forming at the advancing edge of angiogenesis. The deposition of extravasated liposomes in these areas is perivascular and focal, occurring primarily at the roots of capillary sprouts where weak spots (possibly defects or gaps) in the endothelium are believed to occur. Given the geometry of the system, liposomes that enter through such gaps may not be able to penetrate deeply into the tumor interstitium. Liposome penetration may be limited by a range of physical obstacles, including tight cell-cell junctions (often found in highly differentiated epithelial cell tumors), dense connective tissue stroma, small extracellular volume, and high interstitial fluid viscosity (that may be caused by fibrin cross-linking). Ideally all tumor cells, regardless of their proximity to blood vessels or the liposome depots that may form near them, would be exposed to a cytotoxic dose of drug. So, the observation that drug molecules released from focal, perivascular deposits of liposomes are able to penetrate deeply into the tumor mass may be a critical requirement for expression of antitumor activity of PEG-LD.
After many years of development, liposome scientists have come up with a pegylated-coated vesicle in which to carry the potent antitumor agent doxorubicin. These liposomes can circulate for many weeks after injection, being carried by blood vessels to growing tumors. The Stealth liposomes extravasate through gaps into the tumor interstitium and the doxorubicin is then released into the tumor. Free doxorubicin molecules can penetrate deeper into the tumor where they bind to nucleic acids, resulting in tumor cell death.
The ability of PEG-LD to remain in circulation for extended periods of time and to carry doxorubicin to tumor sites contribute to its increased antitumor activity. Another factor in the improved efficacy of PEG-LD over free doxorubicin is its ability to reach higher concentrations in tumor tissue at comparable administered doses.
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