Complete regression of xenografted human carcinomas by a paclitaxel–carboxymethyl dextran conjugate (AZ10992)
Introduction
The anticancer agents paclitaxel (Taxol®) and docetaxel (Taxotere®) are currently the only two US Food and Drug Administration (FDA)-approved representatives of the taxane family. Paclitaxel, an anti-microtubule agent isolated from the trunk bark of the Pacific Yew tree, Taxus brevifolia [1], shows great promise as an anti-neoplastic agent for a variety of human cancers, including breast, ovarian, non-small-cell lung, head and neck [2], [3], [4], [5], [6]. Its unique mechanism of action is related to its ability to promote microtubule assembly and inhibit cell replication in the late G2 or M phase of the cell cycle [7].
A major problem that is associated with the administration of paclitaxel is its low solubility in water (less than 0.01 mg/ml), as well as in most pharmaceutically acceptable solvents. The clinically used formulation of Taxol® contains polyoxyethylated castor oil (Cremophor® EL) and ethanol as excipients. Cremophor® EL has long been considered to be the cause of the hypersensitivity reactions that are observed with paclitaxel infusions [8]. Side effects of Taxol® include nausea, vomiting, diarrhea, mucositis, myelosuppression, cardiotoxicity and neurotoxicity [9], [10]. In an attempt to alleviate the systemic toxicity of paclitaxel, several alternative formulations have been evaluated. Many such forms have entered Phase I or II trials as intravenous injectable anticancer agents – for example, polyglutamate–paclitaxel (XYOTAX, CT-2103; many Phase II/III studies are ongoing); liposome entrapped paclitaxel easy to use (LEP-ETU, PNU-93914; Phase I); N-(2-hydroxypropyl) methacrylamide (HPMA)-copolymer–paclitaxel (PNU-166945; failed at Phase I); polymeric micelle-formulated paclitaxel (Genexol-PM; Phase I); paclitaxel-incorporating micellar nanoparticle formulation (NK105; Phase I). Recently, an albumin-stabilized nanoparticle formulation of paclitaxel (Abraxane, ABI-007) has been approved by the FDA for the treatment of metastatic breast cancer [11].
Polymer–drug conjugates are a subclass of the family of novel drugs and drug-delivery systems that have been defined as ‘polymer therapeutics’ [12]. Polymer therapeutics include polymeric drugs, polymer–drug conjugates, polymer–protein conjugates, polymeric micelles to which the drug is covalently bound, and polymeric non-viral vectors for gene delivery. A number of the first-generation polymer–anticancer-drug conjugates are being tested clinically: HPMA-copolymer–doxorubicin (PK1, FCE28068; Phase II); HPMA-copolymer–doxorubicin–galactosamine (PK2, FCE28069; Phase I/II); HPMA-copolymer–platinate (AP5280; Phase II); HPMA-copolymer–platinate (AP5346; Phase I); and oxidized dextran–doxorubicin (DOX-OXD, AD-70; failed at Phase I). More recently, an increasing number of conjugates containing other anticancer agents – particularly, paclitaxel and camptothecin – are being transferred into the clinic: HPMA-copolymer–camptothecin (MAG-CPT, PNU-166148; failed at Phase I); PEG–camptothecin (PROTECAN; Phase II); polyglutamate–camptothecin (CT-2106; Phase I); and CM dextran–camptothecin derivative (MEN4901/T-0128; Phase I) [13].
These prodrugs are made up of a minimum of three components, i.e. the polymeric polymeric carrier, a biodegradable linker and a bioactive antitumor agent. These macromolecular prodrugs, which are conjugates with prolonged circulation times, target tumors by the ‘enhanced permeability and retention effect’ (EPR). This radically changes the pharmacokinetics of the bound drug. Maeda called the passive targeting phenomenon that he observed, the EPR effect, and attributed it to two factors: the disorganized pathology of tumor vasculature, with its discontinuous endothelium, leading to hyperpermeability to circulating macromolecules, and the lack of effective tumor lymphatic drainage, which leads to subsequent macromolecular accumulation [14], [15]. It is well established that long-circulating macromolecules – including albumin, polymer conjugates, polymeric micelles and liposomes – accumulate passively in solid tumors by the EPR effect, and intravenously administered drug-delivery systems can increase the tumor concentration of antitumor drugs. The polymer–protein conjugate styrene maleic anhydride (SMA)–neocarzinostatin (NCS) – which is called SMANCS – was originally synthesized by Maeda and colleagues, and was subsequently approved in Japan as a treatment for hepatocellular carcinoma. Several conjugates have peptidyl polymer–drug linkers that are amenable to cleavage by lysosomal thiol-dependent proteases. In this case, prodrug activation occurs intracellularly. In contrast, other conjugates that contain an ester link between drug and polymer can release the drug by chemical hydrolysis or esterase degradation extracellularly. Some satisfactory results that have been obtained with the use of macromolecules for the targeted delivery of anticancer agents have generated considerable interest regarding the enhancement of therapeutic efficacy and the reduction of systemic side effects. However, some of the dosage forms have failed, such as oxidized dextran–doxorubicin, probably due to toxicity of the oxidized dextran carrier; and HMPA-copolymer–paclitaxel and HMPA-copolymer–camptothecin, probably due to linkage and/or molecular weight being inadequate.
The concept of a polymer–drug conjugate, combination of suitable polymeric carrier, biodegradable linker and a bioactive antitumor agent, is attractive and could well form the basis of a new generation of anticancer agents. In an attempt to overcome the side effects and to improve the pharmacological profile of paclitaxel, AZ10992 is designed as a polymer–drug conjugate that consists of carboxymethyl (CM) dextran and paclitaxel, linked with the gly–gly–phe–gly (GGFG) linker. CM dextran was chosen as a candidate of the paclitaxel carrier for the following reasons: Dextran is generally recognized to be safe; CM dextran contains a sufficient number of carboxyl groups for drug attachment, which provides sufficient carrying capacity for the drug; and the resulting CM dextran–drug conjugate has a high probability of being water-soluble. We previously reported the preparation of CM dextran and doxorubicin conjugates using a peptide linker. Retention of the conjugate in blood circulation and accumulation of doxorubicin in tumors were both increased by CM dextran with a suitable anionic nature and a molecular weight of 150,000 g/mol [16]. Furthermore, CM dextran–peptide–doxorubicin conjugates containing a GGFG linker were more efficacious in a Walker-256 carcinoma rat model than free doxorubicin or a conjugate with no linker. We also reported the synthesis of CM dextran–paclitaxel conjugates with various amino-acid linkers and the effect of linkers on the in vivo activity [17]. In a tumor distribution study and a tumor growth regression study using intravenous administration of these conjugates in colon26-carcinoma-bearing mice, the amounts of paclitaxel in the tumor and antitumor efficacy were found to correlate. These studies seem to support the use of a CM dextran carrier with a GGFG linker as a way to form an improved drug conjugate.
The purpose of the present study was to test the concept that the rational design of a paclitaxel–polymer conjugate would achieve tumor targeting of the active drug, resulting in improved therapeutic efficacy. To this end, we examined the preclinical profile of the antitumor activity of AZ10992 against colon26 carcinoma in BALB/c mice and a panel of human tumor xenografts in nude mice. The efficacy of AZ10992 was compared with that of non-polymer-bound paclitaxel. In addition, we investigated the plasma and tissue (tumor and non-tumor) pharmacokinetics of both released and polymer-bound paclitaxel after intravenous administration of AZ10992 to mice bearing colon26 carcinoma. These data were also compared with those obtained after dosing of non-polymer-bound paclitaxel to tumor-bearing mice using the same dosage. The results showed potential advantages of AZ10992 that were attributable to the passive tumor targeting.
Section snippets
Chemicals
Paclitaxel (Taxol®) was purchased from Hauser Chemical Research, Inc. (Boulder, CO); Dextran T110 was purchased from Pharmacia Biotech (Uppsala, Sweden); Cremophor® EL and Chymotrypsin were purchased from Sigma Chemical Co. (St. Louis, MO). All other chemicals were of reagent-grade purity or better.
Characterization of AZ10992
The relative molecular weight of AZ10992 was determined using gel permeation chromatography (GPC). A TSK-gel G4000PWXL 300 × 7.8 mm column (TOSOH, Tokyo, Japan) was used, and the column temperature was
In vitro drug release
The in vitro release profiles of paclitaxel from AZ10992 in various conditions – in a buffer, and in plasma and tissue homogenates – are shown in Fig. 2. More than 80% of paclitaxel was liberated from AZ10992 after 24–48 h incubation in plasma or serum. In the buffer, the amount of paclitaxel, as well as 7-epi-paclitaxel released from AZ10992, increased as pH increased. In the buffer (pH 7.4), the amount of released paclitaxel after 24 h incubation was lower than that in plasma or serum. After
Discussion
The objective of this study was to test the concept that the rational design of a paclitaxel–polymer conjugate would achieve the targeting of the tumor with the active drug, resulting in improved therapeutic efficacy. AZ10992 is comprised of paclitaxel that is covalently bound to CM dextran by a GGFG linker. It contains 5.5–6.5 wt.% paclitaxel, molecular weight 150,000 g/mol, linked through the 2′ position – that is, via an ester bond, to the carboxylic acid of polysaccharide. We compared the
Conclusions
AZ10992 was chosen for a paclitaxel tumor-targeting system, with a peptidyl linker GGFG with moderate stability in plasma, and a suitable polysaccharide carrier CM dextran with a molecular weight of 150,000 g/mol. AZ10992 consistently produced regression of tumor xenografts, which are highly refractory to paclitaxel. Altered pharmacokinetic properties of AZ10992 in terms of plasma half-life, tumor targeting and kinetics of paclitaxel release may account for the effectiveness. Results
References (26)
Paclitaxel (Taxol): a novel anticancer chemotherapeutic drug
Mayo Clin. Proc.
(1994)SMANCS and polymer-conjugated macromolecualr drugs: advantages in cancer chemotherapy
Adv. Drug Deliv. Rev.
(1991)- et al.
Mechanisms regulating the recruitment of macrophages into hypoxic areas of tumors and other ischemic tissues
Blood
(2004) - et al.
Macrophage-mediated activation of camptothecin analogue T-2513-carboxymethyl dextran conjugate (T-0128): possible cellular mechanism for antitumor activity
J. Control. Release
(2000) - et al.
Plant antitumor agents. VI. The isolation and structure of taxol, a novel antileukemic and antitumor agent from Taxus brevifolia
J. Am. Chem. Soc.
(1971) - et al.
Cytotoxicity of taxol in vitro against human and rat malignant brain tumors
Cancer Chemother. Pharmacol.
(1994) - et al.
Taxol in malignant melanoma
J. Natl. Cancer Inst. Monographs
(1993) - et al.
Clinical development of taxol
J. Natl. Cancer Inst. Monographs
(1993) - et al.
Taxol (paclitaxel): a novel anti-microtubule agent with remarkable anti-neoplastic activity
Int. J. Clin. Lab. Res.
(1994) - et al.
Cell kill kinetics and cell cycle effects of taxol on human and hamster ovarian cell lines
Cancer Chemother. Pharmacol.
(1993)
Hypersensitivity reactions from taxol
J. Clin. Oncol.
Phase II study and long-term follow-up of patients treated with taxol for advanced ovarian adenocarcinoma
J. Clin. Oncol.
Taxol: Promising new drug of the '90s
Oncol. Nurs. Forum
Cited by (70)
Recent developments in natural biopolymer based drug delivery systems
2023, RSC AdvancesDrug content on anticancer efficacy of self-assembling ketal-linked dextran-paclitaxel conjugates
2023, Journal of Controlled ReleaseComplete regression of xenografted breast tumors by dextran-based dual drug conjugates containing paclitaxel and docosahexaenoic acid
2022, European Journal of Medicinal ChemistryGum-based nanoparticles in cancer therapy
2022, Micro- and Nanoengineered Gum-Based Biomaterials for Drug Delivery and Biomedical ApplicationsBiopolymer-drug conjugates as biomaterials
2021, Tailor-Made and Functionalized Biopolymer Systems: For Drug Delivery and Biomedical Applications