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Clinical Cancer Research 14, 1208, February 15, 2008. doi: 10.1158/1078-0432.CCR-07-0780
© 2008 American Association for Cancer Research
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Cancer Therapy: Preclinical

Irinophore C: A Liposome Formulation of Irinotecan with Substantially Improved Therapeutic Efficacy against a Panel of Human Xenograft Tumors

Euan C. Ramsay1,4, Malathi Anantha1, Jason Zastre1, Marieke Meijs1, Jet Zonderhuis1, Dita Strutt1, Murray S. Webb1, Dawn Waterhouse1,3 and Marcel B. Bally1,2,3

Authors' Affiliations: 1 Department of Advanced Therapeutics, BC Cancer Agency; 2 Department of Pathology and Laboratory Medicine and 3 Faculty of Pharmaceutical Sciences, University of British Columbia, Vancouver, British Columbia, Canada; and 4 School of Pharmacy, University of Tasmania, Hobart, Tasmania, Australia

Requests for reprints: Marcel Bally, Department of Advanced Therapeutics, BC Cancer Agency, 675 West 10th Avenue, Vancouver, British Columbia, Canada V5Z 1L3. Phone: 604-675-8020; Fax: 604-675-8183; E-mail: mbally{at}bccrc.ca.


   Abstract
Top
 Abstract
Materials and Methods
 Results
 Discussion
 References
 
Purpose: To assess the pharmacokinetics, tumor drug accumulation, and therapeutic activity of Irinophore C, a novel liposomal formulation of irinotecan (CPT-11).

Experimental Design: The plasma lactone/carboxy levels of CPT-11 and SN-38 were determined in mice after a single i.v. dose of irinotecan (Camptosar), or Irinophore C, and the plasma t1/2, plasma area under the curve, plasma Cmax, and plasma clearance were calculated. Further, plasma and tumor drug levels were also measured in tumor-bearing mice following Irinophore C treatment. The efficacy of Irinophore C was compared with that of Camptosar in five s.c. human tumor xenografts using single-dose treatment (LS 180), a total of three doses administered at 4-day intervals (H460), or a total of three doses administered at 7-day intervals (Capan-1, PC-3, and HT-29).

Results: Compared with Camptosar, Irinophore C mediated an 8-fold increase in t1/2, a 100-fold increase in Cmax, a 1,000-fold increase in area under the curve, and a 1,000-fold decrease in clearance for the active lactone form of CPT-11. Further, the plasma and tumor SN-38 lactone levels were consistent for at least 48 h post-Irinophore C injection. Camptosar treatment (40 mg/kg) mediated a delay in the time required for tumors to increase to four times their pretreatment size compared with controls (T-C). T-Cs ranged from 2 days (LS 180 model) to 18 days (PC-3 model). Irinophore C (40 mg/kg) engendered T-Cs ranging from 14 days (LS 180 model) to 87 days (Capan-1 model).

Conclusion: Irinophore C improved CPT-11/SN-38 pharmacokinetics, promoted tumor drug accumulation, and increased therapeutic efficacy in a panel of five distinct human tumor xenografts.

Camptothecins mediate their anticancer activity by stabilizing the cleavable complex formed between topoisomerase I and DNA during the S phase of the cell cycle (1); however, this mechanism is dependent on the {alpha}-hydroxy-{delta}-lactone ring of camptothecins (2), which is problematic because physiologic conditions promote the reversible hydrolysis of the lactone ring to yield the inactive carboxylate form (3). Resultantly, research has focused on strategies to stabilize the lactone ring structure, including the use of drug carrier technologies such as hydrogels (4), polymer conjugates (5), microspheres (6), and lipid-based systems (7–12).

The water-soluble camptothecin derivative irinotecan (CPT-11) is an attractive candidate for formulation in a nanocarrier because it has proven clinical activity against colorectal (13) and small-cell lung cancers (14) and may be active in other cancer indications (15). CPT-11 has a complicated pharmacologic profile and is extensively metabolized in vivo to yield a number of derivatives including the potent metabolite SN-38 (16). As with the parent drug, the cytotoxic activity of SN-38 is also dependent on the maintenance of the lactone ring (16).

The physicochemical characteristics of CPT-11 make it amenable to efficient encapsulation in pharmaceutically viable liposome systems. It is already established that CPT-11 can be actively loaded into liposomes via a transmembrane pH gradient (11). The weakly basic drug is added to the outside of liposomes suspended in buffer at pH 7.4, and at this pH a substantial proportion of the drug exists in the neutral form, which can easily permeate the liposomal membrane. On contact with the acidic environment of the aqueous core, CPT-11 is ionized and consequently trapped inside the liposome (17–19). A further benefit of this technology is that the encapsulated CPT-11 exists predominately as the active lactone, which improves therapeutic efficacy (10, 11, 20, 21).

For many anticancer drugs, particularly those that are cell cycle specific, improved drug retention is associated with increases in drug exposure at sites of disease (15, 16). We have recently described a formulation technology that involves the use of entrapped copper ions and a transmembrane pH gradient (acidic inside), which seems to result in significantly improved CPT-11 retention in the liposomes following systemic administration (20). Drugs such as CPT-11 that have protonizable amine groups have been shown to be better retained by liposomes comprising acidic interior buffers (17); further, we have shown that CPT-11 can form a transition metal complex with copper (22). Consequently, following systemic administration, a liposome formulation with a stable internal acidic environment with copper ions mediated substantial improvements in CPT-11 retention, which translated to improved therapeutic activity in a model of colorectal cancer, when compared with carrier systems using pH gradients or copper ion gradients alone. We have named this liposome formulation Irinophore C.

The series of studies described herein investigated the influence of Irinophore C on the pharmacokinetics and tumor accumulation of both the lactone and carboxylate forms of CPT-11 and SN-38. The results will show that Irinophore C mediated a 1,000-fold increase in the plasma CPT-11 lactone area under the curve (AUC) compared with Camptosar and maintained continuous plasma levels of SN-38 lactone for at least 24 h after i.v. administration. Further, CPT-11 lactone and SN-38 lactone accumulated in tumors following Irinophore C treatment. Collectively, the pharmacokinetic improvements and tumor accumulation resulted in substantial gains in therapeutic efficacy in five different xenograft models of human cancer.


   Materials and Methods
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 Abstract
 Materials and Methods
Results
 Discussion
 References
 
Liposome preparation. 1,2-Distearoyl-sn-glycero-phosphocholine and cholesterol (DSPC/Chol; 55:45 mol%; Avanti Polar Lipids) large unilamellar vesicles were prepared as previously described (23). Briefly, lipids were dissolved in chloroform at the required molar ratio, labeled with the nonexchangeable, nonmetabolizable lipid marker 3H-CHE (Perkin-Elmer Life Sciences), and dried to a thin film under a stream of nitrogen gas. Subsequently, the lipid was placed in a high vacuum for ≥3 h to remove any residual solvent. The lipid films were hydrated at 65°C by mixing with 300 mmol/L copper sulfate solution before five cycles of freeze-and-thaw (5 min each, freezing in liquid nitrogen and thawing at 65°C). The multilamellar vesicle suspensions were then extruded 10 times through stacked polycarbonate filters of 0.08- and 0.1-µm pore sizes at 65°C (Extruder, Northern Lipids). The resultant large unilamellar vesicles typically possessed mean vesicular diameters in the range of 110 ± 30 nm as determined using Phase Analysis Light Scattering methods (ZetaPALS, Brookhaven Instruments Corp.). The large unilamellar vesicle external buffer was exchanged using Sephadex G-50 size exclusion chromatography with SHE buffer pH 7.5 (300 mmol/L sucrose, 20 mmol/L HEPES, 15 mmol/L EDTA).

Accumulation of irinotecan into preformed DSPC/Chol liposomes. The divalent metal ionophore A23187 [calcimycin; Sigma; 1 mg/mL (1.9 mmol/L) solution in 100% ethanol] was preincubated with liposomes (0.5 µg/mg lipid) at 60°C for 30 min. Subsequently, irinotecan hydrochloride trihydrate (Camptosar, Pharmacia; BC Cancer Agency Pharmacy) was added to liposomes (~50 mmol/L) at 50°C at a drug-to-lipid ratio of 0.2:1 (mol/mol). Drug uptake was determined after 60-min incubation by separating encapsulated drug from free drug using a Sephadex G-50 column equilibrated with PBS buffer. The excluded fractions, containing the liposomes, were analyzed to determine drug-to-lipid ratios. Lipid concentrations were measured by liquid scintillation counting (Packard 1900TR Liquid Scintillation Analyzer). Irinotecan concentration was determined by measuring absorbance at 370 nm. Briefly, a portion of the sample collected from the column was adjusted to a final volume of 100 µL with PBS. Subsequently, 900 µL of Triton X-100 1% were added and the samples were heated in a water bath at >90°C until the cloud point of the detergent was observed. The samples were then cooled to room temperature and the absorbance was determined against a freshly prepared irinotecan standard curve (Hewlett Packard UV-Vis spectrophotometer, model 8453). For efficacy studies, Irinophore C was concentrated using Amicon Ultra-15 centrifugal Filter Tubes (3,000 x g for 30 min; Millipore) to achieve the desired dose (mg/kg) administered in a volume of 200 µL. The concentrations of lipid and irinotecan present in the final samples were confirmed as described above.

Pharmacokinetic studies. Female BALB/c mice (Taconic; 20-25 g; four per time point) were injected i.v. with a single dose of Camptosar (50 mg/kg irinotecan) or Irinophore C (50 mg/kg irinotecan; 225 mg/kg lipid) and the plasma concentrations of liposomal lipid (3H scintillation counting as described above to measure associated 3H-CHE) and irinotecan were determined over time (11). In addition, female RAG-2M mice (n = 4) with established s.c. human non–small-cell lung cancer cell line (NSCLC) H460 tumors (see "Efficacy studies") were given a single i.v. dose of Irinophore C (20 mg/kg irinotecan; 180 mg/kg lipid) and the plasma levels of drug and lipid were determined as above. High-performance liquid chromatography separation of irinotecan (CPT-11) and SN-38 lactone and carboxylate forms was done using a 250 x 4.6 mm Symmetryshield RP18 5-µm column and Symmetryshield RP18 guard column (Waters). Gradient elution was used with mobile phase A composed of 75 mmol/L ammonium acetate and 7.5 mmol/L tetrabutylammonium bromide adjusted to pH 6.4 with glacial acetic acid (Fisher Scientific) and mobile phase B was acetonitrile. Gradient profile was as follows: time, 0 min: 78% A:22% B; time, 10 min: 64% A:36% B; time, 16 min: 78% A:22% B; time, 20 min: 78% A:22% B. A 10-µL sample was injected onto the column (column temperature, 40°C) and eluted at a flow rate of 1 mL/min. CPT-11 and SN-38 forms were detected using a Waters 2475 multi-wavelength fluorescence detector (Waters) set with time program events of {lambda}ex = 370 nm; {lambda}em = 425 nm between times 0 and 12.5 min for CPT-11 forms, and {lambda}ex = 370 nm; {lambda}em = 535 nm between times 12.5 and 20 min for SN-38 forms. Before injection, all samples were maintained at 4°C to reduce conversion between lactone and carboxylate forms. Standard curves of CPT-11 and SN-38 lactone form were prepared by serial dilutions in a 2:1:1 sodium acetate (100 mmol/L)/methanol/acetonitrile pH 4.0 buffer. For the carboxylate form of CPT-11 and SN-38, serial dilutions were prepared in a 2:1:1 sodium borate (100 mmol/L)/methanol/acetonitrile pH 9.0 buffer. The limit of quantitation for CPT-11 and SN-38 lactone and carboxylate forms was 10 ng/mL. Irinotecan pharmacokinetic parameters were calculated using the noncompartmental analysis function of WinNonLin software version 5.0.1 (Pharsight).

Cell culture. All cell lines were purchased from the American Type Culture Collection and were cultured in the appropriate base media (StemCell) with fetal bovine serum (FBS; Cansera) for up to 10 passages. After 10 passages, new cells were expanded from a frozen stock stored in liquid nitrogen. The human NSCLC H460 was cultured in RPMI 1640 supplemented with 2 mmol/L L-glutamine, 1.5 g/L sodium bicarbonate, 4.5 g/L glucose, 10 mmol/L HEPES, 1.0 mmol/L sodium pyruvate, and 10% FBS. The human colorectal adenocarcinoma cell line LS 180 was cultured in Eagle's MEM with 2 mmol/L L-glutamine and Earle's balanced salt solution, 1.5 g/L sodium bicarbonate, 0.1 mmol/L nonessential amino acids, 1.0 mmol/L sodium pyruvate, and 10% FBS. The human colorectal adenocarcinoma cell line HT-29 was cultured in modified McCoy's 5a medium with 1.5 mmol/L L-glutamine and 2.2 g/L sodium bicarbonate and 10% FBS. The prostate adenocarcinoma cell line PC-3 was cultured in Ham's F12K medium supplemented with 2 mmol/L L-glutamine, 1.5 g/L sodium bicarbonate, and 10% FBS. Capan-1 pancreatic adenocarcinoma cells were cultured in Iscove's modified Dulbecco's medium with 4 mmol/L L-glutamine, 1.5 g/L sodium bicarbonate, and 20% FBS.

Tumor drug accumulation studies. RAG2-M mice (129S6/SvEvTac-Rag2tm1Fwa, Taconic; 20-25 g; three per group) were inoculated s.c. into the center of the lower back with 2 x 106 H460 cells (50 µL). Tumor growth was monitored every Monday, Wednesday, and Friday with calipers and the measured dimensions (in millimeters) were converted to tumor weight (in milligrams) using the following equation: length x (width2) / 2. When the tumors reached ~200 mg, mice were treated with a single dose of Irinophore C (20 mg/kg irinotecan). At the indicated time points, mice were euthanized and plasma and tumors were harvested. Plasma was processed as described above. Tumors were weighed and a 25% homogenate solution was prepared in ice-cold water. CPT-11 and metabolites were extracted by mixing 250 µL of homogenate with 750 µL of ice-cold acetonitrile/methanol (1:1) solution and centrifuging at 14,000 x g for 15 min to precipitate proteins. The supernatant was recovered and a 10-µL aliquot was injected into high-performance liquid chromatography column to quantify the levels of both conformations of CPT-11 and SN-38 using the method outlined above.

Efficacy studies. RAG2-M mice (20-25 g; eight per group) were inoculated s.c. into the center of the lower back with (a) 1 x 106 LS 180 cells (50 µL); (b) 5 x 106 HT-29 cells (50 µL); (c) 2 x 106 H460 cells (50 µL); (d) 5 x 106 PC-3 cells (100 µL); or (e) 2 x 106 Capan-1 cells (50 µL). Once the tumors were established and had reached a mean size of ~25 mg (LS 180), 200 mg (HT-29), 50 mg (H460), 60 mg (PC-3), or 100 mg (Capan-1), mice were treated with Camptosar at doses of 40 or 60 mg/kg (H460 only) or with Irinophore C at doses of 20, 30, or 40 mg/kg, or 60 mg/kg (H460 only), according to the following i.v. dosing schedules: LS 180, single dose; HT-29, each dose administered every 7 days; for a total of 3 doses; H460, each dose administered every 4 days for a total of 3 doses; PC-3, each dose administered every 7 days for a total of 3 doses; Capan-1, each dose administered every 7 days for a total of 3 doses. Tumor growth was monitored with calipers using a schedule governed by the growth characteristics of the tumor model and the measured dimensions (in millimeters) were converted to tumor weight (in milligrams) as described above.

Animal husbandry. Animal studies were conducted in accordance with the Canadian Council on Animal Care Guide, and the University of British Columbia animal care committee approved the animal care protocols. Mice were housed under standard conditions with enrichment and had access to food and water ad libitum. All animals were observed at least twice per day for morbidity, more if deemed necessary during the pretreatment and treatment periods. Signs of ill health were based on body weight loss, change in appetite, and behavioral changes such as altered gait, lethargy, and gross manifestations of stress. Animals were terminated (CO2 asphyxiation) if signs of severe toxicity or tumor-related illness were observed, and a necropsy was done to assess other signs of toxicity.

Statistical analysis. In vivo efficacy data were analyzed using SPSS 13.0 (SPSS, Inc.). The time taken for s.c. tumors to increase 4-fold in relative size [relative tumor growth increase (TGI) = 4] was analyzed using Kaplan-Meier curves for survival analysis. This analysis allows modeling to the time event data (i.e., TGI = 4) in the presence of censored cases (i.e., animals who never reached this end point because of, e.g., tumor ulceration). Treatment and control groups were compared using a log-rank test. Comparisons of the maximum mean percent body weight losses (%BWL) induced following treatments/controls were compared using one-way ANOVA followed by planned contrasts and the Gabriel, Hochberg's GT2, and Games-Howell post hoc tests. The data were normally distributed as determined with the Kolmogorov-Smirnov test. There was no significant difference in variances between groups (Levene's test, P > 0.05).


   Results
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 Abstract
 Materials and Methods
 Results
Discussion
 References
 
A comparison of the pharmacokinetic profiles for Camptosar and Irinophore C. BALB/c mice were injected with a single dose of 50 mg/kg irinotecan (CPT-11), administered as Camptosar or Irinophore C, and the plasma levels of CPT-11 and SN-38, both as lactone and carboxylate conformations, were determined and the results have been summarized in Fig. 1A to D . Camptosar is prepared in acidic solution that favors the active lactone form of CPT-11, and this is reflected in its pharmacokinetic profile (Fig. 1A). The maximum recorded plasma CPT-11 lactone concentration was 10 µg/mL at 5 min postinjection, decreasing by 3 orders of magnitude to 0.01 µg/mL after 8 h (Fig. 1A). In contrast, there was an initial increase in the plasma concentration of CPT-11 carboxylate from 1 µg/mL at the 5-min time point to a maximum of 5 µg/mL after 30 min as the CPT-11 lactone reached equilibrium with the carboxylate conformation under physiologic conditions (Fig. 1A). The elimination profiles of the two forms (lactone and carboxylate) of CPT-11 following Camptosar administration were similar as reflected by their calculated pharmacokinetic parameters (Table 1 ).


Figure 1
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  		Fig. 1. Plasma elimination profiles for CPT-11 and SN-38 species following a single i.v. dose of Camptosar or Irinophore C. Female BALB/c mice were injected i.v. with a single dose of Camptosar (50 mg/kg irinotecan (CPT-11)) or Irinophore C (50 mg/kg CPT-11; 225 mg/kg lipid) and the relative plasma concentrations of CPT-11 lactone ({circ}), CPT-11 carboxylate (bullet), SN-38 lactone ({square}) and SN-38 carboxylate ({blacksquare}) were determined at the indicated time points as described in Materials and Methods. A, Camptosar-Plasma CPT-11 (µg/mL) as a function of time. B, Irinophore C-Plasma CPT-11 (µg/mL) as a function of time. C, Camptosar-Plasma SN-38 (µg/mL) as a function of time. D, Irinophore C-Plasma SN-38 (µg/mL) as a function of time. Points, mean (n = 4); bars, SD. In addition, RAG-2M mice bearing established s.c. human NSCLC H460 xenografts were treated with a single i.v. dose of Irinophore C (20 mg/kg irinotecan; 180 mg/kg lipid) and the plasma concentrations of both forms of CPT-11 and SN-38 were determined. The symbols are as before. E, Irinophore C-Plasma CPT-11 (µg/mL) as a function of time. F, Irinophore C-Plasma SN-38 (µg/mL) as a function of time. Points, mean (n = 4); bars, SD.

 

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  		Table 1. The calculated CPT-11 pharmacokinetic parameters following a single i.v. dose of Camptosar or Irinophore C

 
The plasma concentrations of the two forms of CPT-11 after a single dose of Irinophore C are shown in Fig. 1B. At 1 h postinjection, the CPT-11 lactone concentration was 1,071 µg/mL, and this decreased to 100 µg/mL after 24 h. These levels represent a 10- to 100-fold increase over the maximum recorded concentration achieved following injection of an identical dose of Camptosar. The carrier-mediated increase in CPT-11 levels is highlighted by the pharmacokinetic parameters shown in Table 1 and is best emphasized by the 1,000-fold increase in plasma CPT-11 lactone AUC levels afforded by Irinophore C.

It is assumed that the majority of CPT-11 detected in the plasma following Irinophore C administration is associated with the drug carrier. The pharmacokinetic parameters calculated for Irinophore C CPT-11 carboxylate form supported this belief. For example, the plasma half-life of CPT-11 in the carboxylate form increased from 1 to 9 h, the plasma AUC increased from 9.1 to 55 h µg/mL, and the plasma clearance decreased from 5,475 to 765 mL/kg/h, when compared with results obtained following administration of Camptosar (Table 1). The small proportion of CPT-11 in the carboxylate form following administration of Irinophore C is consistent with the high-performance liquid chromatography and thin liquid chromatography analyses of the formulated product, which exhibits a small amount of CPT-11 in the carboxylate form before injection.

CPT-11 is metabolized by carboxylesterase to yield SN-38, which has been reported to be 100- to 1,000-fold more potent in vitro than CPT-11 (24). In addition, SN-38 also exists in equilibrium between the lactone and carboxylate forms; therefore, the relative plasma levels of the two forms were quantified following Camptosar and Irinophore C i.v. administration. SN-38 lactone and carboxylate forms were both detectable in the plasma 5 min after injection of Camptosar (Fig. 1C). Equilibrium was achieved by the 30-min time point, and the plasma elimination profiles of the two forms were comparable for the remainder of the experiment, with the SN-38 lactone/SN-38 carboxylate ratio consistent at 2:1 (w/w). In contrast, only SN-38 lactone was detectable in the plasma following Irinophore C administration (Fig. 1D). An SN-38 lactone plasma concentration of 2 µg/mL was recorded 1 h postinjection. This level decreased to 1 µg/mL after 4 h and remained constant for the duration of the experiment achieving continuous plasma SN-38 levels over 24 h, which were greater than those observed 10 min after Camptosar administration.

The plasma elimination profiles for both forms of CPT-11 (Fig. 1E) and SN-38 (Fig. 1F) were also quantified following a single dose of Irinophore C 20 mg/kg injected i.v. via the tail vein of RAG-2M mice bearing established s.c. human NSCLC H460 xenografts. The duration of this study was extended to 72 h postinjection and the relative levels of CPT-11 lactone and carboxy for the first 24 h are comparable to that seen in non–tumor-bearing BALB/c mice treated with a higher dose of 50 mg/kg Irinophore C (compare Fig. 1E-B). At 48 h postinjection, the CPT-11 lactone levels had decreased by 2 orders of magnitude (Fig. 1E); in contrast, the concentration of CPT-11 carboxy had remained reasonably constant. Resultantly, the ratio of CPT-11 lactone/CPT-11 carboxy changed from ~1,000:1 over the first 24 h to 10:1 by 72 h. The pharmacokinetic parameters associated with Irinophore C administration to tumor-bearing mice are shown in Table 1 and are comparable to those for BALB/c mice, with a plasma half-life for CPT-11 lactone of 6 h in both models.

The plasma SN-38 levels in the H460 xenograft model following Irinophore C injection (20 mg/kg) are illustrated in Fig. 1F. Consistent with the plasma profile in BALB/c mice (Fig. 1D), SN-38 lactone levels remained steady over 24 h, albeit at concentrations approximately half of that seen with a higher dose of Irinophore C (50 mg/kg), before decreasing by 2 orders of magnitude by 72 h; further, there were no detectable levels of SN-38 carboxy (Fig. 1F). Although SN-38 is known to be highly protein bound (>90%) and the lactone form has a higher affinity thereby forcing equilibrium for unbound plasma SN-38 toward the lactone form (16), it is unclear why SN-38 carboxy was not detected after Irinophore C administration (limit of detection, 10 ng/mL; ref. 25). It may be that SN-38 can be generated when the CPT-11 is still associated with the liposome, thereby facilitating the maintenance of the lactone form (26).

The concentration of CPT-11 and SN-38 in H460 NSCLC tumors following treatment with Irinophore C. Enhanced permeability and retention is a putative mechanism by which nanoparticle drug carrier systems can exert their antitumor effects (27). Particles in the nanometer size range that are normally too large to extravasate from blood vessels are able to passively accumulate in tumor tissue via leaky capillaries associated with the disease. Figure 2 shows the concentrations of CPT-11 and SN-38 present in human NSCLC H460 xenografts grown on the back of RAG-2M mice following a single i.v. dose of Irinophore C (20 mg/kg irinotecan). The drug concentrations have been corrected to account for the blood volume associated with the tumor (28) and, therefore, represent the drug present in the tissue.


Figure 2
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  		Fig. 2. CPT-11 and SN-38 tumor levels in NSCLC H460 xenografts following Irinophore C treatment. RAG2-M mice bearing established s.c. human NSCLC H460 tumors were treated with a single dose of Irinophore C (irinotecan dose of 20 mg/kg) and the tumor lactone/carboxy levels of CPT-11 and SN-38 were measured as described in Materials and Methods. Columns, mean tumor levels (µg/g) collected from three animals, corrected for drug levels associated with the plasma compartment; bars, SD.

 
The concentration CPT-11 lactone (black columns) was the greatest of the species detected over the duration of the experiment, peaking at ~12 µg/g of tumor tissue after 24 h before decreasing to ~5 µg/g of tumor tissue at 48 h. CPT-11 carboxy levels followed the same trend as CPT-11 lactone, albeit at markedly lower levels with a peak of 1 µg/g of tumor tissue detected after 24 h. Because the tumor levels of SN-38 lactone have been corrected for circulating plasma levels, the concentrations shown in Fig. 2 represent local concentrations of SN-38 (29). SN-38 lactone was detectable 1 h postinjection, but levels increased to ~1 µg/g of tumor tissue after 4 h and remained constant for the duration of the experiment, mirroring what was observed in the plasma (see Fig. 1F). SN-38 carboxy was not detected using the experimental methods described in this article.

The therapeutic efficacy of Camptosar and Irinophore C against a panel of five human tumor xenografts. Following i.v. administration of Irinophore C, there was a substantial change in both CPT-11 and SN-38 pharmacokinetics, represented by significant increases in AUC for the lactone forms of these drugs, and the pharmacokinetics suggested that SN-38 levels were maintained over extended time periods; further, CPT-11 lactone and SN-38 lactone were shown to be present in the tumor. The next series of studies aimed to determine whether the altered pharmacokinetic profile of Irinophore C was associated with improvements in therapeutic activity. Camptosar and Irinophore C were used to treat RAG2-M mice bearing established s.c. human tumor xenografts. The results from five different tumor models are summarized in Fig. 3 , where tumor growth is expressed as the median relative TGI as a function of time (days).


Figure 3
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  		Fig. 3. The therapeutic effectiveness of Camptosar and Irinophore C measured against a panel of human tumor xenografts. RAG2-M mice bearing established s.c. human tumor xenografts were treated with saline ({blacktriangleup}), Camptosar 40 mg/kg ({diamondsuit}), Camptosar 60 mg/kg ({blacktriangledown}, H460 only), Irinophore C 20 mg/kg ({circ}), Irinophore C 30 mg/kg ({square}), Irinophore C 40 mg/kg ({lozenge}), and Irinophore C 60 mg/kg ({bigtriangledown}, H460 only). Tumor growth is represented by the median relative TGI and is expressed as a function of time. The number of doses and dosing schedule for each tumor model, as described in Materials and Methods, are indicated by the arrows. Each data point represents n = 8, except for Capan-1 where * indicates that this data point and subsequent data points for this treatment represent n = 7 because there was one durable cure. A, H460 NSCLC; B, LS 180 colorectal adenocarcinoma; C, HT-29 colorectal adenocarcinoma; D, PC-3 prostate carcinoma; E, Capan-1 pancreatic carcinoma.

 
A dose of Camptosar (40 or 60 mg/kg) or Irinophore C (20, 30, 40, or 60 mg/kg) was administered every 4 days for a total of three doses to treat mice with human H460 NSCLC xenografts (Fig. 3A). Camptosar did not result in a significant delay in TGI when compared with the saline-treated controls. Significant improvements (relative to controls and the Camptosar treated animals) were seen following treatment with Irinophore C. This is emphasized by the T-C, which represents the median difference in days for treated tumors (T) compared with control tumors (C) to reach a TGI equal to 4 (Table 2 ). The calculated T-C for the highest Camptosar dose tested (60 mg/kg) was 8 days, significantly lower than the lowest dose of Irinophore C (20 mg/kg), which mediated a T-C of 15 days. The mean %BWL was recorded and used as a marker for toxicity (Table 2). The %BWL associated with Irinophore C 20 mg/kg was 8.4%, which was lower than all treatment groups. There was a trend toward greater %BWL at the highest doses, particularly with Irinophore C 60 mg/kg (%BWL = 18.5%; Table 2), with no significant improvements in therapeutic activity (Fig. 3A; Table 2). For this reason, further efficacy experiments focused on doses equivalent to a maximum of 40 mg/kg CPT-11.


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  		Table 2. The therapeutic effectiveness of Camptosar and Irinophore C against a panel of five human cancer xenografts

 
Figure 3B shows the tumor growth curves for human LS 180 adenocarcinoma xenografts following single-dose treatment with Camptosar or Irinophore C. Consistent with the results seen with the H460 model, all doses of Irinophore C (20, 30, and 40 mg/kg) mediated a greater delay in TGI compared with Camptosar 40 mg/kg, which translated to an ~7-fold increase in T-C if the 40 mg/kg dose data are compared (Table 2). Notably, there was no significant difference in the T-C values between the Irinophore C doses; further, the tumor growth curves were comparable (Fig. 3B). Interestingly, an ~5% BWL was recorded for all treatment groups indicative of Irinophore C having a greater therapeutic index than Camptosar. It was also evident that the body weights had predominately recovered by ~7 days after injection (data not shown); therefore, for the other efficacy models, the dosing interval was increased from 4 days (used to treat H460 tumors) to 7 days.

A treatment schedule of three doses, with each dose administered every 7 days, was sufficient for all doses of Irinophore C to induce tumor regression in a human HT-29 colorectal carcinoma model (Fig. 3C). In contrast, Camptosar 40 mg/kg delayed TGI compared with saline control but did not mediate a regression in tumor size. Resultantly, Irinophore C mediated a 10-fold increase in T-C (32 days for Irinophore C 40 mg/kg compared with 3 days for Camptosar 40 mg/kg; Table 2). Interestingly, there was no significant difference in T-C recorded for the three doses of Irinophore C (Table 2), and this was supported by similar tumor growth curves (Fig. 3C). Analysis of the associated %BWL indicated that administering the treatments at 7-day intervals resulted in similar %BWL observed for single-dose treatment (Table 2; cf. LS 180) and did not result in a cumulative dosing effect that was apparent for H460 (Table 2).

The results for the treatment of RAG2-M mice with established PC-3 prostate carcinoma xenografts followed a pattern similar to that observed for the HT-29 model (i.e., all doses of Irinophore C induced tumor regression whereas Camptosar only delayed tumor growth; Fig. 3D). Further, there were no notable differences in the therapeutic activity of the three Irinophore C doses (Table 2; Fig. 3D), and the %BWL associated with the different treatments were comparable and similar to that recorded for the single-dose treatment of LS 180 (Table 2).

The final model tested was a Capan-1 pancreatic xenograft and the results are shown in Fig. 3E. This was the only model within the panel of human tumor xenografts where there was a marked difference in therapeutic efficacy associated with the dose of Irinophore C. Clearly, all doses of Irinophore C were substantially more active than Camptosar 40 mg/kg (Fig. 3E). Moreover, the highest dose of Irinophore C (40 mg/kg) was able to cure one mouse and also mediated a T-C (87 days) that was significantly greater than those recorded for the 20 and 30 mg/kg doses, as well as being 15-fold higher than the T-C for the equivalent dose of Camptosar (6 days; Table 2). Consistent with the previous models using the dosing schedule of three doses administered every 7 days, the %BWL were comparable to those observed following a single dose (Table 2).


   Discussion
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 Abstract
 Materials and Methods
 Results
 Discussion
References
 
Liposomes represent the preeminent nanoscale drug delivery technology for the i.v. administration of cytotoxic drugs (30), with three anthracycline preparations currently licensed for clinical use (31–33) and many other liposomal formulations of antineoplastic drugs in preclinical or clinical trials (12, 34–37). The beneficial effects of liposomes have been attributed to decreased toxicity, increased drug stability, and improved biodistribution of the encapsulated drugs (38).

The clinical effect of CPT-11 on overall patient survival and quality of life has been significant (39) despite the physiologic instability of the active lactone forms of the drug and its potent metabolite, SN-38 (3). Consequently, CPT-11 is an attractive candidate for formulation with lipid nanocarriers. Burke and Gao (7) first showed that lipid-based carrier formulations could stabilize the lactone ring of topotecan; subsequently, the stabilization of CPT-11 lactone, following i.v. administration, by liposome encapsulation has been associated with improved therapeutic activity (10, 11, 20). Irinophore C is a novel liposome formulation of CPT-11 in which drug retention following systemic administration was greatly improved by entrapping the drug in an acidic copper ion environment (40).

Following a single i.v. bolus injection, Irinophore C mediated a substantial increase in the CPT-11 lactone plasma half-life, maximum plasma concentration (Cmax), and plasma AUC and a substantial decrease in plasma clearance, when compared with an equivalent dose of Camptosar (Table 1). Likewise, Irinophore C altered the pharmacokinetic profile for CPT-11 carboxylate (Table 1; Fig. 1A and B). Presumably, this indicates that the majority of both forms of CPT-11 detected in the plasma are associated with the carrier system, which would explain why the high CPT-11 Cmax levels following i.v. administration of Irinophore C did not precipitate a "cholinergic syndrome." This toxicity is attributed to CPT-11–mediated acetylcholinesterase inhibition (41) and limits the maximum single i.v. dose of Camptosar that RAG2-M mice can tolerate to 80 mg/kg. In contrast, Irinophore C is well tolerated at this dose in RAG-2M mice and has been administered to the plasma carboxylesterase–deficient murine strain aES1e/J (42) at single doses up to 350 mg/kg with no immediate or long-term toxicities.5 Drummond et al. (10) reported similar results for a liposomal irinotecan formulation in which multivalent anionic trapping agents were used to increases retention of the drug.

Protracted dosing schedules would be expected to be beneficial for S-phase active drugs like CPT-11 (15, 16). Clinical studies have shown that the maximum CPT-11 dose intensity achievable with long-term continuous infusion is actually two to three times lower than that observed with short-term infusions; however, the SN-38 plasma AUC levels were comparable (15, 16). This implies that higher CPT-11 doses may saturate the hepatic carboxylesterases that are predominately responsible for the generation of SN-38. Further, there is evidence to suggest that optimal anticancer activity is dependent on the maintenance of a threshold level of exposure to SN-38 (43). Furman et al. (44) reported that a prolonged CPT-11 dosing schedule optimized in xenograft models to maintain a minimum threshold plasma concentration of SN-38 could be mimicked in children with a comparable relationship between SN-38 levels and response. The long half-life of CPT-11 release from the carrier (44.4 h; ref. 40) could prevent saturation of the carboxylesterase enzymes in the liver, enabling Irinophore C to mediate a constant plasma SN-38 lactone level over the duration of the experiment (Fig. 1D and F). Indeed, the inherent accumulation of liposomes in the liver might act as a drug depot where the macrophage disruption of the carrier could slowly free CPT-11 for enzymatic conversion (45). If the lowest dose of Irinophore C (20 mg/kg) still maintained the plasma SN-38 levels above the critical threshold (Fig. 1F), this could explain the lack of dose-response effect observed for the majority of the xenograft models tested (Fig. 3).

The mechanism of tumor growth delay, or shrinkage, observed in the five xenograft models following Irinophore C treatment remains to be determined. Camptothecins have been reported to have antiangiogenic effects (46), and it may be that maintenance of a threshold plasma level of SN-38 by Irinophore C could mimic metronomic dosing, in which the anticancer action is primarily through disruption of the tumor vasculature. However, despite SN-38 being reported as up to 1,000 times more potent than CPT-11 in vitro (24), it is difficult to estimate the contribution of each compound to the overall cytotoxic effect in vivo (47). There have been in vitro survival studies using HT-29 cells (48) and H460 cells (25) that have attributed the observed cytotoxicity solely to CPT-11. Analysis of H460 tumor levels of CPT-11 (Fig. 2) showed that the lactone form predominates, suggesting either that CPT-11 is still encapsulated within the liposome or, alternatively, that the lower pH of the tumor microenvironment is limiting the conversion of bioavailable CPT-11 lactone to the carboxy form (49). SN-38 lactone levels within the tumor, after accounting for the presence of the active metabolite in the plasma, could be due to CPT-11 lactone that leaked from regionally localized liposomes, or it could be from the plasma compartment. The latter could be a result of drug release from liposomes within the plasma compartment or following localization in other tissues such as the liver. However, it is also reasonable to speculate that tumor accumulation of liposomes via the enhanced permeability and retention effect and subsequent leakage of CPT-11 could also contribute to the anticancer effect of Irinophore C. We are currently investigating whether the activity of Irinophore C is mediated by a dual-action mechanism encompassing antivascular and direct tumor cell cytotoxic actions. Further, ongoing studies in our laboratory are also assessing the therapeutic efficacy of Irinophore C against a systemic melanoma model grown in aES1e/J mice. These mice are deficient in plasma carboxylesterase and are believed to offer a better model of SN-38 metabolism in humans. It is hoped that these experiments will provide a more mechanistic basis for the improved therapeutic effects of Irinophore C.

Obviously, further clinical development of Irinophore C will also require a better understanding of the toxicity of this new formulation. The principal dose-limiting toxicity of irinotecan is delayed diarrhea, which is attributed to the high intestinal concentrations of SN-38 excreted directly via the biliary canal, or as the result of β-glucuronidase cleavage of SN-38G to SN-38 by the gut flora (50). Ongoing studies in our laboratory are investigating the effect of CPT-11 encapsulation on the principal toxicities associated with irinotecan therapy.

In conclusion, Irinophore C is a novel liposomal formulation of irinotecan, which substantially increases the plasma levels of CPT-11 lactone and mediates a prolonged plasma and tumor exposure to SN-38 lactone. This favorable pharmacokinetic profile translated into superior therapeutic activity compared with Camptosar when tested against a panel of five human cancer xenografts. These results would support the further development of Irinophore C for future clinical trials.


   Footnotes
 
Grant support: Canadian Institute for Health Research (M.B. Bally, D. Waterhouse, E.C. Ramsay, and M.S. Webb).

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

5 Unpublished observations. Back

Received 4/ 4/07; revised 10/ 2/07; accepted 11/26/07.


   References
Top
 Abstract
 Materials and Methods
 Results
 Discussion
 References
 

  1. Hsiang YH, Lihou MG, Liu LF. Arrest of replication forks by drug-stabilized topoisomerase I-DNA cleavable complexes as a mechanism of cell killing by camptothecin. Cancer Res 1989;49:5077–82.[Abstract/Free Full Text]
  2. Giovanella BC, Harris N, Mendoza J, Cao Z, Liehr J, Stehlin JS. Dependence of anticancer activity of camptothecins on maintaining their lactone function. Ann N Y Acad Sci 2000;922:27–35.[Medline]
  3. Burke TG. Chemistry of the camptothecins in the bloodstream. Drug stabilization and optimization of activity. Ann N Y Acad Sci 1996;803:29–31.[Medline]
  4. Lalloo A, Chao P, Hu P, Stein S, Sinko PJ. Pharmacokinetic and pharmacodynamic evaluation of a novel in situ forming poly(ethylene glycol)-based hydrogel for the controlled delivery of the camptothecins. J Control Release 2006;112:333–42.[CrossRef][Medline]
  5. Vicent MJ, Duncan R. Polymer conjugates: nanosized medicines for treating cancer. Trends Biotechnol 2006;24:39–47.[CrossRef][Medline]
  6. Shenderova A, Burke TG, Schwendeman SP. The acidic microclimate in poly(lactide-co-glycolide) microspheres stabilizes camptothecins. Pharm Res 1999;16:241–8.[CrossRef][Medline]
  7. Burke TG, Gao X. Stabilization of topotecan in low pH liposomes composed of distearoylphosphatidylcholine. J Pharm Sci 1994;83:967–9.[CrossRef][Medline]
  8. Sadzuka Y, Hirotsu S, Hirota S. Effective irinotecan (CPT-11)-containing liposomes: intraliposomal conversion to the active metabolite SN-38. Jpn J Cancer Res 1999;90:226–32.[CrossRef]
  9. Chou TH, Chen SC, Chu IM. Effect of composition on the stability of liposomal irinotecan prepared by a pH gradient method. J Biosci Bioeng 2003;95:405–8.[Medline]
  10. Drummond DC, Noble CO, Guo Z, Hong K, Park JW, Kirpotin DB. Development of a highly active nanoliposomal irinotecan using a novel intraliposomal stabilization strategy. Cancer Res 2006;66:3271–7.[Abstract/Free Full Text]
  11. Messerer CL, Ramsay EC, Waterhouse D, et al. Liposomal irinotecan: formulation development and therapeutic assessment in murine xenograft models of colorectal cancer. Clin Cancer Res 2004;10:6638–49.[Abstract/Free Full Text]
  12. Pal A, Khan S, Wang YF, et al. Preclinical safety, pharmacokinetics and antitumor efficacy profile of liposome-entrapped SN-38 formulation. Anticancer Res 2005;25:331–41.[Abstract/Free Full Text]
  13. Saltz LB, Cox JV, Blanke C, et al. Irinotecan plus fluorouracil and leucovorin for metastatic colorectal cancer. Irinotecan Study Group. N Engl J Med 2000;343:905–14.[Abstract/Free Full Text]
  14. Noda K, Nishiwaki Y, Kawahara M, et al. Irinotecan plus cisplatin compared with etoposide plus cisplatin for extensive small-cell lung cancer. N Engl J Med 2002;346:85–91.[Abstract/Free Full Text]
  15. Garcia-Carbonero R, Supko JG. Current perspectives on the clinical experience, pharmacology, and continued development of the camptothecins. Clin Cancer Res 2002;8:641–61.[Abstract/Free Full Text]
  16. Mathijssen RH, van Alphen RJ, Verweij J, et al. Clinical pharmacokinetics and metabolism of irinotecan (CPT-11). Clin Cancer Res 2001;7:2182–94.[Abstract/Free Full Text]
  17. Cullis PR, Hope MJ, Bally MB, Madden TD, Mayer LD, Fenske DB. Influence of pH gradients on the transbilayer transport of drugs, lipids, peptides and metal ions into large unilamellar vesicles. Biochim Biophys Acta 1997;1331:187–211.[Medline]
  18. Haran G, Cohen R, Bar LK, Barenholz Y. Transmembrane ammonium sulfate gradients in liposomes produce efficient and stable entrapment of amphipathic weak bases. Biochim Biophys Acta 1993;1151:201–15.[Medline]
  19. Facchini FM, Spiro SG. Chemotherapy in small-cell lung cancer: is more better? In: Brambilla C, Brambilla E, editors. Lung tumors: fundamental biology and clinical management. New York: Marcel Dekker, Inc.; 1999. p. 611–30.
  20. Mayer LD, Harasym TO, Tardi PG, et al. Ratiometric dosing of anticancer drug combinations: controlling drug ratios after systemic administration regulates therapeutic activity in tumor-bearing mice. Mol Cancer Ther 2006;5:1854–63.[Abstract/Free Full Text]
  21. Sadzuka Y, Hirotsu S, Hirota S. Effect of liposomalization on the antitumor activity, side-effects and tissue distribution of CPT-11. Cancer Lett 1998;127:99–106.[CrossRef][Medline]
  22. Ramsay E, Alnajim J, Anantha M, et al. Transition metal-mediated liposomal encapsulation of irinotecan (CPT-11) stabilizes the drug in the therapeutically active lactone conformation. Pharm Res 2006;23:2799–808.[CrossRef][Medline]
  23. Hope MJ, Bally M, Mayer L, Janoff AS, Cullis P. Generation of multilamellar and unilamellar phospholipid vesicles. Chem Phys Lipids 1986;40:89–107.[CrossRef]
  24. Lavelle F, Bissery MC, Andre S, Roquet F, Riou JF. Preclinical evaluation of CPT-11 and its active metabolite SN-38. Semin Oncol 1996;23:11–20.[Medline]
  25. Zastre J, Anantha M, Ramsay E, Bally M. Irinotecan-cisplatin interactions assessed in cell-based screening assays: cytotoxicity, drug accumulation and DNA adduct formation in an NSCLC cell line. Cancer Chemother Pharmacol 2007;60:91–102.[CrossRef][Medline]
  26. Sadzuka Y. Effective prodrug liposome and conversion to active metabolite. Curr Drug Metab 2000;1:31–48.[CrossRef][Medline]
  27. Maeda H, Wu J, Sawa T, Matsumura Y, Hori K. Tumor vascular permeability and the EPR effect in macromolecular therapeutics: a review. J Control Release 2000;65:271–84.[CrossRef][Medline]
  28. Bally MB, Mayer LD, Hope MJ, Nayar R. Pharmacodynamics of liposomal drug carriers: methodological considerations. In: Gregoriadis G, editor. Liposome technology. London: CRC Press; 1993. p. 27–41.
  29. van Ark-Otte J, Kedde MA, van der Vijgh WJ, et al. Determinants of CPT-11 and SN-38 activities in human lung cancer cells. Br J Cancer 1998;77:2171–6.[Medline]
  30. Allen TM, Cullis PR. Drug delivery systems: entering the mainstream. Science 2004;303:1818–22.[Abstract/Free Full Text]
  31. Northfelt DW, Dezube BJ, Thommes JA, et al. Pegylated-liposomal doxorubicin versus doxorubicin, bleomycin, and vincristine in the treatment of AIDS-related Kaposi's sarcoma: results of a randomized phase III clinical trial. J Clin Oncol 1998;16:2445–51.[Abstract]
  32. Harris L, Batist G, Belt R, et al. Liposome-encapsulated doxorubicin compared with conventional doxorubicin in a randomized multicenter trial as first-line therapy of metastatic breast carcinoma. Cancer 2002;94:25–36.[CrossRef][Medline]
  33. Gill PS, Wernz J, Scadden DT, et al. Randomized phase III trial of liposomal daunorubicin versus doxorubicin, bleomycin, and vincristine in AIDS-related Kaposi's sarcoma. J Clin Oncol 1996;14:2353–64.[Abstract]
  34. Semple SC, Leone R, Wang J, et al. Optimization and characterization of a sphingomyelin/cholesterol liposome formulation of vinorelbine with promising antitumor activity. J Pharm Sci 2005;94:1024–38.[CrossRef][Medline]
  35. Seiden MV, Muggia F, Astrow A, et al. A phase II study of liposomal lurtotecan (OSI-211) in patients with topotecan resistant ovarian cancer. Gynecol Oncol 2004;93:229–32.[CrossRef][Medline]
  36. Thomas DA, Sarris AH, Cortes J, et al. Phase II study of sphingosomal vincristine in patients with recurrent or refractory adult acute lymphocytic leukemia. Cancer 2006;106:120–7.[CrossRef][Medline]
  37. Stathopoulos GP, Boulikas T, Kourvetaris A, Stathopoulos J. Liposomal oxaliplatin in the treatment of advanced cancer: a phase I study. Anticancer Res 2006;26:1489–93.[Abstract/Free Full Text]
  38. Harrington KJ, Syrigos KN, Vile RG. Liposomally targeted cytotoxic drugs for the treatment of cancer. J Pharm Pharmacol 2002;54:1573–600.[CrossRef][Medline]
  39. Pessino A, Sobrero A. Optimal treatment of metastatic colorectal cancer. Expert Rev Anticancer Ther 2006;6:801–12.[CrossRef][Medline]
  40. Ramsay E, Alnajim J, Anantha M, et al. A novel liposomal irinotecan formulation with significant anti-tumor activity: use of the divalent cation ionophore A23187 and copper-containing liposomes to improve drug retention. Eur J Pharm Biopharm. 2007 Sep 2 [Epub ahead of print].
  41. Tobin P, Rivory L, Clarke S. Inhibition of acetylcholinesterase in patients receiving irinotecan (camptothecin-11). Clin Pharmacol Ther 2004;76:505–6; author reply 07–8.[CrossRef][Medline]
  42. Morton CL, Iacono L, Hyatt JL, et al. Activation and antitumor activity of CPT-11 in plasma esterase-deficient mice. Cancer Chemother Pharmacol 2005;56:629–36.[CrossRef][Medline]
  43. Jung LL, Zamboni WC. Cellular, pharmacokinetic, and pharmacodynamic aspects of response to camptothecins: can we improve it? Drug Resist Updat 2001;4:273–88.[CrossRef][Medline]
  44. Furman WL, Stewart CF, Poquette CA, et al. Direct translation of a protracted irinotecan schedule from a xenograft model to a phase I trial in children. J Clin Oncol 1999;17:1815–24.[Abstract/Free Full Text]
  45. Storm G, Steerenberg PA, Emmen F, van Borssum Waalkes M, Crommelin DJ. Release of doxorubicin from peritoneal macrophages exposed in vivo to doxorubicin-containing liposomes. Biochim Biophys Acta 1988;965:136–45.[Medline]
  46. Clements MK, Jones CB, Cumming M, Daoud SS. Antiangiogenic potential of camptothecin and topotecan. Cancer Chemother Pharmacol 1999;44:411–6.[CrossRef][Medline]
  47. Rivory LP. Irinotecan. In: Adams VR, Burke TG, editors. Camptothecins in cancer therapy. Totowa: Humana Press Inc.; 2005. p. 229–62.
  48. Pavillard V, Agostini C, Richard S, Charasson V, Montaudon D, Robert J. Determinants of the cytotoxicity of irinotecan in two human colorectal tumor cell lines. Cancer Chemother Pharmacol 2002;49:329–35.[CrossRef][Medline]
  49. Gerweck LE, Vijayappa S, Kozin S. Tumor pH controls the in vivo efficacy of weak acid and base chemotherapeutics. Mol Cancer Ther 2006;5:1275–9.[Abstract/Free Full Text]
  50. Yang X, Hu Z, Chan SY, et al. Novel Agents that Potentially Inhibit Irinotecan-Induced Diarrhea. Curr Med Chem 2005;12:1343–58.[CrossRef][Medline]



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J. H.E. Baker, J. Lam, A. H. Kyle, J. Sy, T. Oliver, S. J. Co, W. H. Dragowska, E. Ramsay, M. Anantha, T. J. Ruth, et al.
Irinophore C, a Novel Nanoformulation of Irinotecan, Alters Tumor Vascular Function and Enhances the Distribution of 5-Fluorouracil and Doxorubicin
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