Mutagenic effect of borocaptate sodium and boronophenylalanine in neutron capture therapy

PURPOSE: To investigate the mutagenic effect in boron neutron capture therapy (BNCT), Chinese hamster ovary cells were incubated with 10 B-enriched borocaptate sodium (BSH) or para-boronophenylalanine (BPA) before exposure to thermal neutrons, and the occurrence of mutations at the hypoxanthine-guanine phosphoribosyl transferase (HPRT) locus was measured. METHODS AND MATERIALS: BSH or BPA was added to cells 20 h or 2 h before irradiation and removed before irradiation. Cells were irradiated with thermal neutrons. The biologic end point of cell survival was measured by colony formation assay. The mutagenicity was calculated from the mutation frequency at the HPRT locus. RESULTS: The mutagenicity of BSH and BPA was similar to that of 10B boric acid when the cells were irradiated with neutrons at an isosurvival dose after 2-h preincubation. Preincubation with BSH for 20 h, compared with preincubation for just 2 h, had no effect on either cytotoxicity or mutagenicity in BNCT. However, with BPA, 20-h preincubation, compared with 2-h preincubation, caused an increase in the cell killing effect, but a decrease in the mutagenic effect of the BNCT. CONCLUSION: After 20-h incubation, BPA was less mutagenic than BSH. The mutagenic study of electroporated BPA or BSH revealed a reduced mutagenicity. These results suggest that the retention of these boron compounds in the cells causes a more accurate assault on the cell and lessens the chance of misrepair after neutron irradiation.     

Effect of electroporation on cell killing by boron neutron capture therapy using borocaptate sodium (10B-BSH).

The cell membrane permeability of 10B-enriched borocaptate sodium (BSH) and the extent to which BSH is accumulated in cells are controversial. To elucidate these points and to enhance the accumulation of BSH in cells, the effect of electroporation on boron neutron capture therapy (BNCT) using BSH was investigated. The first group of SCCVII tumor cells was incubated in culture medium with 10B-BSH or 10B-enriched boric acid, and exposed to neutrons from the heavy water facility of the Kyoto University Reactor. More than 99% of neutrons were thermal neutrons at flux base. The second group was pretreated with electroporation in combination with 10B-BSH, and thereafter the cells were irradiated with neutrons. The cell-killing effect of BNCT was measured by colony formation assay. The surviving cell fraction decreased exponentially with neutron fluence, and addition of BSH significantly enhanced the cell-killing effect of NCT depending on 10B concentration and the preincubation time of cells in the BSH-containing culture medium. The electroporation of cells with BSH markedly enhanced the BNCT effect in comparison with that obtained with preincubation alone. The effect of BSH-BNCT with electroporation was almost equal to that of BNCT using 10B-boric acid at the same 10B concentration. The effect of BNCT on cells pretreated with BSH and electroporation was not reduced by repeated washing of the cells before neutron irradiation. Decrease of the effect of BSH-BNCT plus electroporation with increase in the waiting time between the electroporation and the neutron irradiation could be explained in terms of the extent of cell growth during that time. These data suggest that BSH penetrates the cells slowly and remains after washing. Electroporation can introduce BSH into the cells very efficiently, and BSH thus introduced stays in the cells and is not lost in spite of the intensive washing of the cells. Therefore, if electroporation is applied to tumors after BSH injection, 10B would remain in the tumors but be cleared from normal tissues, and selective accumulation of 10B in tumors will be achieved after an appropriate waiting time.     

The combined effect of boronophenylalanine and borocaptate in boron neutron capture therapy for SCCVII tumors in mice

PURPOSE: To increase the effect of boron neutron capture therapy (BNCT) on tumors in vivo, the combined effects of para-boronophenylalanine (BPA) and borocaptate sodium (BSH) were investigated. METHODS AND MATERIALS: 10B-enriched BPA and BSH were administered to C3H/He mice bearing SCCVII tumors by intragastric and intravenous injections, respectively. The colony formation and tumor control assays were employed for investigating antitumor effects of BNCT. The extent of homogeneity of tumor cell killing effect was examined by the distribution of frequencies of binuclear cells (BNC) producing a certain number of micronuclei (0,1,2,--,> or =5) to total number of BNC and by the comparison between surviving cell fraction (SF) in colony formation assay and the normal nuclear division fraction (NNDF) at first mitosis following BNCT. RESULTS: The relationships between SF and radiation dose in Gy (D) at around 10 ppm of 10B in tumors were as follow: -InSF = -0.101 + 0.648 Gy(-1) x D, 0.0606+0.435 Gy(-1) x D, and -0.0155 + 0.342 Gy(-1) x D for BPA, BPA + BSH, and BSH, respectively. In tumor control assay, BPA was also more effective than BSH, but the difference of effectiveness significantly decreased: 1.9 times more effective in colony assay vs. 1.2 times in tumor control assay. The most effective treatment to achieve tumor cure was BNCT using BPA + BSH, and it was 1.9 times more effective than BSH-BNCT. In BSH-BNCT, NNDF decreased exponentially with radiation dose and was equal to SF. However, NNDF following BPA-BNCT showed a biphasic decrease with radiation dose, and SF was much lower than NNDF. In the combination of BPA and BSH, the discrepancy between NNDF and SF decreased in comparison with BPA-BNCT. The distribution of frequency of BNC with a certain number of micronuclei to total BNC was very close to Poisson distribution in BSH-BNCT tumors; however, it deviated from the Poisson in BPA-BNCT tumors. In combination with BPA and BSH, the distribution showed an intermediate pattern. These findings indicate that BSH distributes homogeneously with a heterogeneous distribution of BPA in tumors, and the heterogeneous effect of BPA-BNCT was improved by the combination of two boron compounds. CONCLUSION: The heterogeneous cell killing effect of BPA-BNCT was improved by the combination of BSH, and increased tumor control rates. Therefore, this combination may improve clinical outcome of BNCT although the effects on normal tissues have to be examined before clinical application.     

Pharmacokinetics of sodium borocaptate: a critical assessment of dosing paradigms for boron neutron capture therapy

Summary The pharmacokinetics of sodium borocaptate (BSH), a drug that has been used clinically for boron neutron capture therapy (BNCT) of malignant brain tumors, have been characterized by measuring boron concentrations by direct current plasma-atomic emission spectroscopy (DCP-AES) in a group of 23 patients with high-grade gliomas. The disposition of BSH following intravenous (i.v.) infusion, which was determined by measuring plasma boron concentrations by DCP-AES, was consistent with a three-compartment open model with zero-order input and first-order elimination from the central compartment. Boron disposition was linear over the dose range of 26.5–88.2 mg BSH/kg body weight (b.w.), corresponding to 15–50 mg boron/kg b.w. Mean total body boron plasma clearance was 14.4 ± 3.5 ml/min and the harmonic mean half-lives (range) were 0.6 (0.3–3.7), 6.5 (4.8–10.1) and 77.8 (49.6–172.0) h for theα, β, andγ disposition phases, respectively. Using an empirically determined plasma: blood boron concentration ratio of 1.3 ± 0.2, the calculated total body boron blood clearance was 18.5 ± 4.5 ml/min. In order to develop a model for selecting the optimum dosing paradigm, a pharmacokinetic correlation was established between the boron content of normal brain, solid tumor, and infiltrating tumor to the shallow tissue pharmacokinetic compartment (C₂). Based on our model, it was concluded that although multiple i.v. infusions of BSH might increase absolute tumor boron concentrations, they will not improve the tumor: plasma boron concentration ratios over those attainable by a single i.v. infusion. The results from our study are confirmatory of those previously reported by others when blood sampling has been carried out for a sufficient period of time to adequately characterize the pharmacokinetics.     

Tumor-specific targeting of sodium borocaptate (BSH) to malignant glioma by transferrin-PEG liposomes: a modality for boron neutron capture therapy

Object Boron neutron capture therapy (BNCT) requires selective delivery of a high concentration of boron-10 (¹⁰B) to tumor tissue. To improve a drug delivery in BNCT, we devised transferrin-conjugated polyethylene-glycol liposome encapsulating sodium borocaptate (TF-PEG-BSH). Methods ¹⁰B concentrations of U87Δ human glioma cells from three boron delivery systems (BDS) (bare BSH, PEG-BSH, and TF-PEG-BSH) were analyzed in vitro by use of inductively coupled plasma-atomic emission spectrometry (ICP-AES). A colony-forming assay (CFA) was performed using this cell line with the three BDS and neutron irradiation. Subcellular localization of BSH in tumor cells was analyzed in vitro by immunocytochemistry using monoclonal antibodies raised for BSH. Brain tumor models were made and the ¹⁰B concentrations of the tumor, blood, and normal brain from each BDS were analyzed in vivo by use of ICP-AES. The tumor-to-blood and the tumor-to-normal brain ratios were evaluated. BNCT for the brain tumor models was performed and survival was analyzed. Results In CFA, TF-PEG-BSH showed the most prominent effects by neutron irradiation among the three BDS. TF-PEG-BSH showed highly selective and highly efficient ¹⁰B delivery in tumor tissue. The survival rate in the treatment experiment was best in the TF-PEG-BSH group. Immunocytochemically, TF-PEG-BSH delivered BSH efficiently not only into the cytoplasm but also into the nucleus. Conclusion TF-PEG-BSH is a potent BDS for BNCT not only in terms of delivering a high concentration of ¹⁰B into tumor tissue, but also the selective delivery of ¹⁰B into the tumor cells.     

Role of p53 mutation in the effect of boron neutron capture therapy on oral squamous cell carcinoma

Background

To evaluate diagnosis, management and outcome of breast cancer (BC) occurring after irradiation for Hodgkin's lymphoma (HL).

Methods

39 cases of BC in 28 HL survivors were retrospectively reviewed. 21 patients were included in a case-control analysis.

Results

The median age at diagnosis of HL and BC was 25.3 and 45.3 years, respectively. The median interval to develop BC was 16.1 years. Eleven women (39.2%) had bilateral disease. Mode of detection of the index breast cancers was by mammographic screening in 17 patients (60.7%), palpable lump in 8 patients (28.6%), clinical examination in two patients (7.1%), and unknown in one patient (3.6%). Case-control analysis showed that histological features and prognosis of BC after HL were similar to those of primary BC, however, for BC after HL, mastectomy was the predominant surgery (P =.001) and adjuvant radiotherapy and anthracycline-based chemotherapy were less frequently used as compared to primary BC (P <.001 and.003, respectively).

Conclusion

The previous history of HL does not appear to be a poor prognostic factor for BC occurring thereafter.     

Optimal timing of neutron irradiation for boron neutron capture therapy after intravenous infusion of sodium borocaptate in patients with glioblastoma

PURPOSE: A cooperative study in Europe and Japan was conducted to determine the pharmacokinetics and boron uptake of sodium borocaptate (BSH: Na(2)B(12)H(11)SH), which has been introduced clinically as a boron carrier for boron neutron capture therapy in patients with glioblastoma. METHODS AND MATERIALS: Data from 56 patients with glioblastoma who received BSH intravenous infusion were retrospectively reviewed. The pharmacokinetics were evaluated in 50 patients, and boron uptake was investigated in 47 patients. Patients received BSH doses between 12 and 100 mg/kg of body weight. For the evaluation, the infused boron dose was scaled linearly to 100 mg/kg BSH. RESULTS: In BSH pharmacokinetics, the average value for total body clearance, distribution volume of steady state, and mean residence time was 3.6 +/- 1.5 L/h, 223.3 +/- 160.7 L, and 68.0 +/- 52.5 h, respectively. The average values of the boron concentration in tumor adjusted to 100 mg/kg BSH, the boron concentration in blood adjusted to 100 mg/kg BSH, and the tumor/blood boron concentration ratio were 37.1 +/- 35.8 ppm, 35.2 +/- 41.8 ppm, and 1.53 +/- 1.43, respectively. A good correlation was found between the logarithmic value of T(adj) and the interval from BSH infusion to tumor tissue sampling. About 12-19 h after infusion, the actual values for T(adj) and tumor/blood boron concentration ratio were 46.2 +/- 36.0 ppm and 1.70 +/- 1.06, respectively. The dose ratio between tumor and healthy tissue peaked in the same interval. CONCLUSION: For boron neutron capture therapy using BSH administered by intravenous infusion, this work confirms that neutron irradiation is optimal around 12-19 h after the infusion is started.     

First attempt of boron neutron capture therapy (BNCT) for hepatocellular carcinoma

A 60-year-old man with multiple hepatocellular carcinomas (HCCs) was enrolled as the first patient in a pilot study for treating multiple liver tumors with boron neutron capture therapy (BNCT). Because of compromised liver function, the multiple tumors in the right liver lobe were treated with BNCT and those in the left lobe with hepatic arterial chemoembolization. The feasibility and clinical outcome of this first case was assessed. During the treatment and follow-up period, no adverse effect as a result of BNCT was observed except for temporary temperature elevation to 38.3 degrees C, and the AST and ALT being higher than 200 IU/l. For 1 month, the tumors treated with BNCT remained stable in size. The BNCT-treated tumors showed regrowth 3.5 months after BNCT and the patient died of liver dysfunction caused by progression of HCC 10 months after BNCT. The feasibility of BNCT for HCC is confirmed in this first case.     

Dosimetric study of boron neutron capture therapy with borocaptate sodium (BSH)/lipiodol emulsion (BSH/lipiodol-BNCT) for treatment of multiple liver tumors

PURPOSE: We performed a computational study to investigate the feasibility of borocaptate sodium (BSH)/lipiodol-boron neutron capture therapy (BSH/lipiodol-BNCT) for multiple liver tumors using Simulation Environment for Radiotherapy Applications (SERA), a currently available BNCT treatment planning system. METHODS AND MATERIALS: Three treatment plans for BSH/lipiodol-BNCT using two or three epithermal neutron beams in one fraction were generated for 4 patients with multiple liver tumors using the SERA system. The (10)B concentrations in the tumor and the liver assumed in the study were 197.3 and 15.3 ppm, respectively; and were obtained from experimental studies in animals. The therapeutic gain factors for the liver tumors, defined as the minimum dose to the tumor/maximum dose to the liver, and the inhomogeneity index of the thermal neutron fluence for the whole of the liver, defined as the maximum neutron fluence - minimum neutron fluence/mean neutron fluence, were evaluated in each plan. RESULTS: Three epithermal neutron beams incident on the anterior, posterior, and right side of the patient can deliver the most homogeneous distribution of thermal neutron fluence to the whole of the liver and provide the greatest therapeutic gain factors for tumors in the right lobe and approximately equal therapeutic gain factors for tumors in the left lobe, compared with the two opposed (anterior-posterior) and two orthogonal (anterior-right) beams. CONCLUSIONS: From a dosimetric viewpoint, the BSH/lipiodol-BNCT treatment plan using three epithermal neutron beams is the most suitable for the treatment of multiple liver tumors.     

Boron neutron capture therapy of brain tumors: enhanced survival and cure following blood-brain barrier disruption and intracarotid injection of sodium borocaptate and boronophenylalanine

PURPOSE: Boronophenylalanine (BPA) and sodium borocaptate (Na(2)B(12)H(11)SH or BSH) have been used clinically for boron neutron capture therapy (BNCT) of high-grade gliomas. These drugs appear to concentrate in tumors by different mechanisms and may target different subpopulations of glioma cells. The purpose of the present study was to determine if the efficacy of BNCT could be further improved in F98-glioma-bearing rats by administering both boron compounds together and by improving their delivery by means of intracarotid (i.c.) injection with or without blood-brain barrier disruption (BBB-D). METHODS AND MATERIALS: For biodistribution studies, 10(5) F98 glioma cells were implanted stereotactically into the brains of syngeneic Fischer rats. Eleven to 13 days later animals were injected intravenously (i.v.) with BPA at doses of either 250 or 500 mg/kg body weight (b.w.) in combination with BSH at doses of either 30 or 60 mg/kg b.w. or i.c. with or without BBB-D, which was accomplished by i.c. infusion of a hyperosmotic (25%) solution of mannitol. For BNCT studies, 10(3) F98 glioma cells were implanted intracerebrally, and 14 days later animals were transported to the Brookhaven National Laboratory (BNL). They received BPA (250 mg/kg b.w.) in combination with BSH (30 mg/kg b.w. ) by i.v. or i.c. injection with or without BBB-D, and 2.5 hours later they were irradiated with a collimated beam of thermal neutrons at the BNL Medical Research Reactor. RESULTS: The mean tumor boron concentration +/- standard deviation (SD) at 2.5 hours after i. c. injection of BPA (250 mg/kg b.w.) and BSH (30 mg/kg b.w.) was 56. 3 +/- 37.8 microgram/g with BBB-D compared to 20.8 +/- 3.9 microgram/g without BBB-D and 11.2 +/- 1.8 microgram/g after i.v. injection. Doubling the dose of BPA and BSH produced a twofold increase in tumor boron concentrations, but also concomitant increases in normal brain and blood levels, which could have adverse effects. For this reason, the lower boron dose was selected for BNCT studies. The median survival time was 25 days for untreated control rats, 29 days for irradiated controls, 42 days for rats that received BPA and BSH i.v., 53 days following i.c. injection, and 72 days following i.c. injection + BBB-D with subsets of long-term survivors and/or cured animals in the latter two groups. No histopathologic evidence of residual tumor was seen in the brains of cured animals. CONCLUSIONS: The combination of BPA and BSH, administered i.c. with BBB-D, yielded a 25% cure rate for the heretofore incurable F98 rat glioma with minimal late radiation-induced brain damage. These results demonstrate that using a combination of boron agents and optimizing their delivery can dramatically improve the efficacy of BNCT in glioma-bearing rats.     

The combined effect of electroporation and borocaptate in boron neutron capture therapy for murine solid tumors.

10 B-Enriched borocaptate (BSH) was administered intraperitoneally to SCCVII tumor-bearing C3H / He mice. Electroporation (EP) was conducted by using a tweezers-type electrode. The (10) B contents in tumors were measured by prompt gamma-ray spectrometry. The colony formation assay was applied to investigate the antitumor effects of boron neutron capture therapy (BNCT) and thereby to estimate the intratumor localization of BSH. The (10) B concentrations in tumors decreased with time following BSH administration, falling to 5.4(0. 1) ppm at 3 h, whereas EP treatment (3 repetitions) 15 min after BSH injection delayed the clearance of BSH from tumors, and the (10) B level remained at 19.4(0.9) ppm at 3 h. The effect of BNCT increased with the (10) B concentration in tumors, and the combination with EP showed a remarkably large cell killing effect even at 3 h after BSH injection. The effect of BNCT, i.e., slope coefficient of the cell survival curve of tumors, without EP was proportional to tumor (10) B level (r = 0.982), and that of BSH-BNCT combined with EP lay close to the same correlation line. However, tumors subjected to EP after BSH injection did not show high radiosensitivity when irradiated after conversion to a single cell suspension by enzymatic digestion. This indicates that the increase of the BNCT effect by EP was a consequence of enclosure of BSH in the interstitial space of tumor tissue and not within tumor cells. This is different from a previous in vitro study. The combination of EP and BNCT may be clinically useful, if a procedure to limit EP to the tumor region becomes available or if an alternative similar method is employed.     

The hamster cheek pouch as a model of oral cancer for boron neutron capture therapy studies: selective delivery of boron by boronophenylalanine.

Herein we propose and validate the hamster cheek pouch model of oral cancer for boron neutron capture therapy (BNCT) studies. This model serves to explore new applications of the technique, study the biology and radiobiology of BNCT, and assess the uptake of boron compounds and response of tumor, precancerous tissue, and clinically relevant normal tissues. These issues are central to evaluating and improving the therapeutic gain of BNCT. The success of BNCT is dependent on the absolute amount of boron in the tumor, and the tumor:blood and tumor:normal tissue boron concentration ratios. Within this context, biodistribution studies are pivotal. Tumors were induced in the hamsters with a carcinogenesis protocol that uses dimethyl-1,2-benzanthracene and mimics spontaneous tumor development in human oral mucosa. The animals were then used for biodistribution and pharmacokinetic studies of boronophenylalanine (BPA). Blood, tumor, precancerous pouch tissue surrounding tumor, normal pouch tissue, tongue, skin, cheek mucosa, palate mucosa, liver, and spleen, were sampled at 0-12 h after administration of 300 mg BPA/kg. The data reveal selective uptake of BPA by tumor tissue and, to a lesser degree, by precancerous tissue. Mean tumor boron concentration was 36.9 +/- 17.5 ppm at 3.5 h and the mean boron ratios were 2.4:1 for tumor:normal pouch tissue and 3.2:1 for tumor:blood. Higher doses of BPA (600 and 1200 mg BPA/kg) increased tumor uptake. Potentially therapeutic absolute boron concentrations, and tumor:normal tissue and tumor:blood ratios can be achieved in the hamster oral cancer model using BPA as the delivery agent.     

Experimental application of boron neutron capture therapy to undifferentiated thyroid carcinoma

PURPOSE: Undifferentiated thyroid carcinoma (UTC) lacks an effective treatment. Boron neutron capture therapy (BNCT) is based on the nuclear reaction (10)B(n,alpha) (7)Li. These particles destroy the tumor locally due to their high linear energy transfer (LET). Mice transplanted with the human cell line of UTC ARO have a selective uptake of (10)B-borophenylalanine (BPA). The complete BNCT was performed to explore its possible application. METHODS AND MATERIALS: Mice were distributed into four groups: (1) no treatment; (2) neutron beam alone; (3) 350 mg/kg body weight (b.w.) BPA plus irradiation; (4) 600 mg/kg b.w. BPA plus irradiation. Follow-up was performed by measurement of tumor volume, histologic analysis, and assessment of DNA damage using the comet assay. RESULTS: The tumor continued to grow in Groups 1 and 2. In Group 3, a slow-down of tumor growth was observed in all mice, and a complete stop was observed in 100% of mice of Group 4. Complete disappearance of the tumor was observed in 50% of the mice that had an initial tumor volume of less than 50 mm(3) (Groups 3 and 4). DNA damage showed a progressive increase from Group 1 through 4. CONCLUSION: These data show, for the first time, that UTC is amenable to treatment by BNCT.     

Intra-arterial administration of sodium borocaptate (BSH)/lipiodol emulsion delivers B-10 to liver tumors highly selectively for boron neutron capture therapy: experimental studies in the rat liver model

PURPOSE: Boron neutron capture therapy (BNCT) is particle radiotherapy with alpha ((4)He) particle and recoiled lithium nucleus ((7)Li) derived from a reaction of boron ((10)B) and thermal neutron. We investigated applying BNCT to malignant liver tumors. The purpose of the present study was to reveal the efficacy for administration of emulsion of a boron compound (sodium borocaptate; BSH) and lipiodol via a hepatic artery using a rat liver tumor model. METHODS AND MATERIALS: Rat liver tumors were developed by direct injection of Walker 256 cells into the liver parenchyma. BSH (75 mg/kg)/lipiodol (0.3 mL/kg) emulsion was administered via the hepatic artery. Boron concentrations in the tumors, liver, and blood were measured at 1, 6, and 12 h after administration. Neutron capture radiography (NCR) was taken to confirm the selective accumulation of (10)B in the liver tumors. RESULTS: Boron concentrations in the liver tumors and the tumor/liver (T/L) boron concentration ratio at 1, 6, and 12 h after administration of BSH/lipiodol emulsion (concentration: T/L ratio) were 479.2 ppm: 4.0, 197.3 ppm: 14.9, and 96.5 ppm: 6.6, respectively. Highly selective irradiation was clearly demonstrated by the NCR images. CONCLUSIONS: Intra-arterial administration of BSH/lipiodol emulsion is effective method for delivering high concentration of (10)B selectively to the liver tumors.     

Combined use of sodium borocaptate and buthionine sulfoximine in boron neutron capture therapy enhanced tissue boron uptake and delayed tumor growth in a rat subcutaneous tumor model

We have previously reported that buthionine sulfoximine (BSO) enhances sodium borocaptate (BSH) uptake by down regulating glutathione (GSH) synthesis in cultured cells. This study investigated the influence of BSO on tissue BSH uptake in vivo and the efficacy of BSH-BSO-mediated boron neutron capture therapy (BNCT) on tumor growth using a Fisher-344 rat subcutaneous tumor model. With BSO supplementation, boron uptake in subcutaneous tumor, blood, skin, muscle, liver, and kidney was significantly enhanced and maintained for 12h. Tumor growth was significantly delayed by using BSO. With further improvement in experimental conditions, radiation exposure time, together with radiation damage to normal tissues, could be reduced.     

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

PURPOSE: To evaluate GB-10-encapsulating transferrin (TF)-pendant-type polyethyleneglycol (PEG) liposomes as tumor-targeting (10)B-carriers for boron neutron capture therapy. METHODS AND MATERIALS: A free mercaptoundecahydrododecaborate-(10)B (BSH) or decahydrodecaborate-(10)B (GB-10) solution, bare liposomes, PEG liposomes, or TF-PEG liposomes were injected into SCC VII tumor-bearing mice, and (10)B concentrations in the tumors and normal tissues were measured by gamma-ray spectrometry. Meanwhile, tumor-bearing mice were continuously given 5-bromo-2'-deoxyuridine (BrdU) to label all intratumor proliferating cells, then injected with these (10)B-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 (10)B 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 (10)B-carriers were significantly greater than BSH-containing (10)B-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 (10)B-carriers.     

Selective targeting of boronophenylalanine to melanoma in BALB/c mice for neutron capture therapy

Melanoma cells actively accumulate aromatic amino acids for use as precursors in the synthesis of the pigment melanin. Using the Harding-Passey melanoma carried s.c. in BALB/c mice, we have demonstrated that p-boronophenylalanine (BPA) is taken up by melanoma tissue to a much greater extent than by normal tissues. Following a single i.p. injection, or a series of injections given over 1 h, the accumulation of boron in melanoma was found to be transient, reaching a maximum approximately 6 h postinjection. The concentrations of boron achieved in tumor ranged from 9-33 micrograms/g, and are within the range estimated to be necessary for successful application of the nuclear reaction 10B(n,alpha)7Li for neutron capture therapy. Boron concentrations in tumor and tissues were determined using either a prompt-gamma spectroscopic technique or by quantitative neutron capture radiography using whole-body sections. Distribution studies with the resolved stereoisomers of BPA indicated that the L isomer is preferentially accumulated in the melanoma compared to the D isomer. The L isomer of BPA was shown to be targeted to actively dividing tumor cells by simultaneously comparing the boron and [3H]thymidine distribution in tumor. Under conditions which selectively deliver high concentrations of boron to Harding-Passey melanomas in BALB/c mice, BPA did not deliver useful concentrations of boron to a mammary adenocarcinoma in Hale-Stoner mice. These results, along with the selectivity of the Harding-Passey melanoma for the L isomer of BPA, are consistent with our working hypothesis that BPA is actively transported into the melanomas as an analogue of natural melanin precursors.     

Boron microlocalization in oral mucosal tissue: implications for boron neutron capture therapy

Clinical studies of the treatment of glioma and cutaneous melanoma using boron neutron capture therapy (BNCT) are currently taking place in the USA, Europe and Japan. New BNCT clinical facilities are under construction in Finland, Sweden, England and California. The observation of transient acute effects in the oral mucosa of a number of glioma patients involved in the American clinical trials, suggests that radiation damage of the oral mucosa could be a potential complication in future BNCT clinical protocols, involving higher doses and larger irradiation field sizes. The present investigation is the first to use a high resolution surface analytical technique to relate the microdistribution of boron-10 (10B) in the oral mucosa to the biological effectiveness of the 10B(n,alpha)7Li neutron capture reaction in this tissue. The two boron delivery agents used clinically in Europe/Japan and the USA, borocaptate sodium (BSH) and p-boronophenylalanine (BPA), respectively, were evaluated using a rat ventral tongue model. 10B concentrations in various regions of the tongue mucosa were estimated using ion microscopy. In the epithelium, levels of 10B were appreciably lower after the administration of BSH than was the case after BPA. The epithelium:blood 10B partition ratios were 0.2:1 and 1:1 for BSH and BPA respectively. The 10B content of the lamina propria was higher than that measured in the epithelium for both BSH and BPA. The difference was most marked for BSH, where 10B levels were a factor of six higher in the lamina propria than in the epithelium. The concentration of 10B was also measured in blood vessel walls where relatively low levels of accumulation of BSH, as compared with BPA, was demonstrated in blood vessel endothelial cells and muscle. Vessel wall:blood 10B partition ratios were 0.3:1 and 0.9:1 for BSH and BPA respectively. Evaluation of tongue mucosal response (ulceration) to BNC irradiation indicated a considerably reduced radiation sensitivity using BSH as the boron delivery agent relative to BPA. The compound biological effectiveness (CBE) factor for BSH was estimated at 0.29 +/- 0.02. This compares with a previously published CBE factor for BPA of 4.87 +/- 0.16. It was concluded that variations in the microdistribution profile of 10B, using the two boron delivery agents, had a significant effect on the response of oral mucosa to BNC irradiation. From a clinical perspective, based on the findings of the present study, it is probable that potential radiation-induced oral mucositis will be restricted to BNCT protocols involving BPA. However, a thorough high resolution analysis of 10B microdistribution in human oral mucosal tissue, using a technique such as ion microscopy, is a prerequisite for the use of experimentally derived CBE factors in clinical BNCT.     

Enhanced survival of glioma bearing rats following boron neutron capture therapy with blood-brain barrier disruption and intracarotid injection of boronophenylalanine

Boronophenylalanine (BPA) has been used for boron neutron capture therapy (BNCT) of brain tumors in both experimental animals and humans. The purpose of the present study was to determine if the efficacy of BNCT could be enhanced by means of intracarotid (i.c.) injection of BPA with or without blood-brain barrier disruption (BBB-D) and neutron irradiation using a rat brain tumor model. For biodistribution studies, F98 glioma cells were implanted stereotactically into the brains of Fischer rats, and12 days later BBB-D was carried out by i.c. infusion of 25% mannitol (1.373 mOsmol/ml), followed immediately by i.c. administration of 300, 500 or 800 mg of BPA/kg body weight (b.w.). At the 500 mg dose a fourfold increase in tumor boron concentration (94.5 g/g) was seen at 2.5 hours after BBB-D, compared to 20.8 g/g in i.v. injected animals. The best composite tumor to normal tissue ratios were observed at 2.5 hours after BBB-D, at which time the tumor: blood (T: Bl) ratio was10.9, and the tumor: brain (T: Br) ratio was 7.5, compared to 3.2 and 5.0 respectively for i.v. injected rats. In contrast, animals that had received i.c. BPA without BBB-D had T: Bl and T: Br ratios of 8.5 and 5.9, respectively, and the tumor boron concentration was 42.7g/g. For therapy experiments, initiated 14 days after intracerebral implantation of F98 glioma cells, 500 mg/kg b.w. of BPA were administered i.v. or i.c. with or without BBB-D, and the animals were irradiated 2.5 hourslater at the Brookhaven Medical Research Reactor with a collimated beam of thermal neutrons delivered to the head. The mean survival time for untreated control rats was 24 ± 3 days, 30 ± 2 days for irradiated controls, 37 ± 3 days for those receiving i.v. BPA, 52 ± 15 days for rats receiving i.c. BPA without BBB-D, and 95 ± 95 days for BBB-D followed by i.c. BPA and BNCT. The latter group had a 246% increase in life span (ILS) compared to untreated controls and a 124% ILS compared to that of i.v. injected animals. These survival data are the best ever obtained with the F98 glioma model and suggest that i.c. administration of BPA with or without BBB-D may be useful as a means to increase the efficacy of BNCT.     

A critical assessment of boron target compounds for boron neutron capture therapy

Summary Boron neutron capture therapy (BNCT) has undergone dramatic developments since its inception by Locher in 1936 and the development of nuclear energy during World War II. The ensuing Cold War spawned the entirely new field of polyhedral borane chemistry, rapid advances in nuclear reactor technology and a corresponding increase in the number to reactors potentially available for BNCT. This effort has been largely oriented toward the eradication of glioblastoma multiforme (GBM) and melanoma with reduced interest in other types of malignancies. The design and synthesis of boron-10 target compounds needed for BNCT was not channeled to those types of compounds specifically required for GBM or melanoma. Consequently, a number of potentially useful boron agents are known which have not been biologically evaluated beyond a cursory examination and only three boron-10 enriched target species are approved for human use following their Investigational New Drug classification by the US Food and Drug Administration; BSH, BPA and GB-10. All ongoing clinical trials with GBM and melanoma are necessarily conducted with one of these three species and most often with BPA. The further development of BNCT is presently stalled by the absence of strong support for advanced compound evaluation and compound discovery driven by recent advances in biology and chemistry. A rigorous demonstration of BNCT efficacy surpassing that of currently available protocols has yet to be achieved. This article discusses the past history of compound development, contemporary problems such as compound classification and those problems which impede future advances. The latter include means for biological evaluation of new (and existing) boron target candidates at all stages of their development and the large-scale synthesis of boron target species for clinical trials and beyond. The future of BNCT is bright if latitude is given to the choice of clinical disease to be treated and if a recognized study demonstrating improved efficacy is completed. Eventually, BNCT in some form will be commercialized.