The potential of transferrin-pendant-type polyethyleneglycol liposomes encapsulating decahydrodecaborate-10B (GB-10) as 10B-carriers for boron neutron capture therapy

doi:10.1016/j.ijrobp.2006.08.028

International Journal of Radiation OncologyBiologyPhysics

Volume 66, Issue 5, 1 December 2006, Pages 1515-1522

https://doi.org/10.1016/j.ijrobp.2006.08.028 Get rights and content

Purpose: To evaluate GB-10-encapsulating transferrin (TF)-pendant-type polyethyleneglycol (PEG) liposomes as tumor-targeting ¹⁰B-carriers for boron neutron capture therapy.

Methods and Materials: A free mercaptoundecahydrododecaborate-¹⁰B (BSH) or decahydrodecaborate-¹⁰B (GB-10) solution, bare liposomes, PEG liposomes, or TF-PEG liposomes were injected into SCC VII tumor-bearing mice, and ¹⁰B concentrations in the tumors and normal tissues were measured by γ-ray spectrometry. Meanwhile, tumor-bearing mice were continuously given 5-bromo-2′-deoxyuridine (BrdU) to label all intratumor proliferating cells, then injected with these ¹⁰B-carriers containing BSH or GB-10 in the same manner. Right after thermal neutron irradiation, the response of quiescent (Q) cells was assessed in terms of the micronucleus frequency using immunofluorescence staining for BrdU. The frequency in the total tumor cells was determined from the BrdU nontreated tumors.

Results: Transferrin-PEG liposomes showed a prolonged retention in blood circulation, low uptake by reticuloendothelial system, and the most enhanced accumulation of ¹⁰B in solid tumors. In general, the enhancing effects were significantly greater in total cells than Q cells. In both cells, the enhancing effects of GB-10–containing ¹⁰B-carriers were significantly greater than BSH-containing ¹⁰B-carriers, whether loaded in free solution or liposomes. In both cells, whether BSH or GB-10 was employed, the greatest enhancing effect was observed with TF-PEG liposomes followed in decreasing order by PEG liposomes, bare liposomes, and free BSH or GB-10 solution. In Q cells, the decrease was remarkable between PEG and bare liposomes.

Conclusions: In terms of biodistribution characteristics and tumor cell–killing effect as a whole, including Q cells, GB-10 TF-PEG liposomes were regarded as promising ¹⁰B-carriers.

Introduction

A neutron capture reaction in boron [¹⁰B (n,α)⁷ Li] is, in principle, very effective in destroying tumors, providing that a sufficient amount of ¹⁰B can be accumulated in the target tumor and a sufficient number of very low energy thermal neutrons can be delivered there. The two particles generated in this reaction carry a high linear energy transfer and have a range of roughly the diameter of one or two tumor cells. It is theoretically possible to kill tumor cells without affecting adjacent healthy cells, if ¹⁰B atoms can be selectively accumulated in the interstitial space of tumor tissue or intracellular space of tumor cells. Thus, the success in boron neutron capture therapy (BNCT) requires the selective delivery of large amounts of ¹⁰B to malignant cells. At the same time, the ¹⁰B concentration in the surrounding normal tissue should be kept low to minimize damage to the normal tissue (1).

Various approaches have been employed for the delivery of ¹⁰B to tumors, including the use of macromolecules such as monoclonal antibodies, epidermal growth factor, and dextran conjugates, and microparticles such as liposomes, low-density lipoprotein complexes, and microcapsules (2). In particular, the use of liposomes could be a promising approach because they can carry large quantities of ¹⁰B leading to selective localization in tumors (3). Further, the inclusion of polyethylene glycol (PEG) was reported to significantly reduce the uptake by the reticuloendothelial system (RES) of liposomes, resulting in their prolonged circulation (4). Recently, a new type of target-sensitive liposome bearing PEG, so-called pendant-type PEG immunoliposomes, in which antibodies or specific ligands are coupled to the extremities of surface-grafted PEG chains, has been reported (5). In particular, transferrin-coupling pendant-type PEG liposomes (TF-PEG liposomes) were demonstrated to extravasate effectively into solid tumors and internalize into tumor cells (5). TF-PEG liposomes showed a prolonged circulation and low RES uptake in tumor-bearing mice, resulting in enhanced extravasation of the liposomes into solid tumors.

To make liposomes containing enough ¹⁰B, mercaptoundecahydrododecaborate-¹⁰B (sodium borocaptate-¹⁰B, BSH) (Fig. 1), which has been widely used for clinical trials of BNCT for the treatment of glioblastoma multiforme (1), or its alternative ¹⁰B-compound, decahydrodecaborate-¹⁰B (GB-10) (Fig. 1) (6), was employed as an encapsulated ¹⁰B-carrier in the liposomes. GB-10 forms the anion (B_10H10)⁻² in aqueous solution. It is manufactured by the oxidation of decaborane and has no special handling or storage requirements. GB-10 is a largely diffusible agent that does not traverse the intact blood–brain barrier (6). It was shown to be nontoxic in dogs (7) and proposed as a boron agent for BNCT and BNCT-enhanced fast neutron therapy (8).

In this study, we examined the potential of liposomes for the selective delivery of therapeutic quantities of ¹⁰B to tumors. TF-PEG liposomes and PEG liposomes encapsulating BSH or GB-10 were prepared and their tissue distributions in tumor-bearing mice after intravenous injection were compared with those of conventional liposomes (bare liposomes) and a free BSH or GB-10 solution. Based on the findings in biodistribution studies, we selected a suitable dosage of each ¹⁰B-carrier and a time point for starting thermal neutron irradiation for BNCT. We then examined the effects of these ¹⁰B-carriers on total (proliferating [P] + quiescent [Q]) and Q tumor cell populations in combination with thermal neutron irradiation, in terms of the surviving fraction (SF) and the micronucleus (MN) frequency, using our own method for selectively detecting the Q cell response to DNA-damaging treatment (9).

Section snippets

Preparation of TF-pendant-type PEG liposomes encapsulating BSH or GB-10

Bare liposomes and PEG liposomes were prepared from DSPC (COATSOME MC-8080) and CH (Wako, Osaka, Japan) and from DSPC, CH, DSPE-PEG, and DSPE-PEG-COOH, respectively. DSPE (COATSOME ME-8080)-PEG and DSPE-PEG-COOH were synthesized as previously described (10). Small unilamellar vesicles of these two liposomes were prepared according to the reverse-phase evaporation method (11). Three hundred milligrams of lipid were dissolved in 4 mL of chloroform/diethyl ether mixture (1:1 v/v). Two milliliters

Results

Overall, the MN frequencies at 0 Gy were larger for Q cells than for the total cells. Notably, when any ¹⁰B-carrier other than bare liposomes was used, these differences were significant (p < 0.05) (Table 1). Further, the SFs and the MN frequencies at 0 Gy were significantly lower and higher, respectively, using GB-10 than without any ¹⁰B-carrier and using BSH (p < 0.05). In both total and Q-cell populations, whether BSH or GB-10 was employed, the SFs decreased and the MN frequencies increased

Discussion

In general, liposomes can carry a large amount of ¹⁰B leading to selective localization in tumors (3). Actually, the blood levels of ¹⁰B remained high for a long time, and the administration of liposomal BSH or GB-10 increased the accumulation of ¹⁰B in solid tumors compared with the use of free BSH or GB-10 solution only. Further, the blood concentrations of ¹⁰B were much higher using TF-PEG liposomes and PEG liposomes than using ¹⁰B-loaded bare liposomes. This confirmed the effectiveness of

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: Supported, in part, by a Grant-in-aid for Scientific Research (C) (18591380) from the Japan Society for the Promotion of Science.

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Biology contributionThe potential of transferrin-pendant-type polyethyleneglycol liposomes encapsulating decahydrodecaborate-10B (GB-10) as 10B-carriers for boron neutron capture therapy

Introduction

Section snippets

Preparation of TF-pendant-type PEG liposomes encapsulating BSH or GB-10

Results

Discussion

Int J Pharm

Biochim Biophys Acta

Nucl Instrum Methods

Nucl Instr Meth Phys Res A

Adv Drug Deliv Rev

Int J Radiat Oncol Biol Phys

Semin Radiat Oncol

Boron neutron capture therapy of cancer: Current status and future prospects

Clin Cancer Res

Boron containing macromolecules as nanovehicles as delivery agents for neutron capture therapy

Anti-Cancer Agents Med Chem

Inhibition of human pancreatic cancer growth in nude mice by boron neutron capture therapy

Br J Cancer

Biology contribution
The potential of transferrin-pendant-type polyethyleneglycol liposomes encapsulating decahydrodecaborate-¹⁰B (GB-10) as ¹⁰B-carriers for boron neutron capture therapy