The high mortality and morbidity of Head and Neck cancer patients and the toxicity associated to oncological treatments suggest the need for more selective and non-toxic therapies. BNCT (Boron Neutron Capture Therapy) is based on the capture reaction between boron-10, selectively targeted to tumor tissue, and a thermal neutron. In this study, an oral cancer and precancer model was used to study the therapeutic effect and radiotoxicity of BNCT mediated by BPA (boronphenylalanine) [BPA-BNCT] at short and medium-term follow-up. Oligo-Fucoidan, an extract of Laminaria japonica brown seaweed, was evaluated as a potentially beneficial adjunct to BPA-BNCT. Hamsters were chemically cancerized over 8 weeks. Tumor bearing hamsters were assigned to: (CONTROL) cancerized, sham-irradiated group; BPA-BNCT 3 Gy absorbed dose to precancerous tissue; BPA-BNCT 3 Gy + Oligo-Fucoidan (200 mg/kg/day, for 16 days). Neutron irradiation was performed at the RA-3 Nuclear Reactor 3 h post administration of BPA (15.5 mg 10B/kg). Boron biodistribution and microdistribution studies were performed in BPA and BPA + Oligo-Fucoidan groups. Oligo-Fucoidan did not reduce either the incidence of moderate/severe mucositis or reduced the percentage of animals with new tumors in precancerous tissue. Oligo-Fucoidan did enhance BPA-BNCT-induced tumor response at one month (67–94

BNCT is a radiotherapeutic modality of high LET energy for different solid tumors as colorectal carcinoma (CRC). Although clinical outcomes show advantages of BNCT, tumor recurrence remains a common challenge. Previously we have described the double strand DNA damage response (DDR) produced by BNCT. TGF-β/SMAD pathway has been involved in maintaining genomic integrity. The aim of these studies was to evaluate the activation of the TGF-β/SMAD pathway, its interaction with the DDR pathway and the possible use of LY2109761 (Ly), a specific inhibitor of TGF-β receptor, as a radiosensitizer for BNCT. Six groups were performed in a human colon adenocarcinoma (HT29) cell line: NCT (neutrons), BNCT (boronophenylalanine plus neutrons), Control and the same three groups with the addition of Ly. The results showed an activation of the TGF-β/SMAD cascade with an increase in the genomic expression of TGF-β, SMAD7 and ATR (p < 0.001) at 2 h post neutron treatments compared to the Control group. A significant decrease in the expression of TGF-β receptor type I, SMAD7 and ATR for BNCT plus Ly was observed. Furthermore, it was demonstrated a decrease in tumor survival as a function of the total absorbed physical dose for all the treatments, being significantly higher in the groups treated with Ly at 1 and 3 Gy. On the other hand, a lower number of Ki67+ cells with the addition of Ly was found. Conclusion: The activation of the TGF-β/SMAD pathway and its interaction with the DNA repair via through ATR transductor was demonstrated. LY2109761 could act as a radiosensitizer for BNCT.

Background and Aims: Boron Neutron Capture Therapy (BNCT) is a tumor selective particle radiotherapy that combines preferential accumulation of 10B compounds and neutron irradiation. BPA (boronophenylalanine) has been used in clinical trials. LAT1 is responsible for its uptake in tumor cells and is upregulated in Head and Neck Cancer (HNC). Our group studied BPA biodistribution and BPA/BNCT in the hamster cheek pouch oral cancer model. In this model, exophytic tumors are surrounded by a precancerous tissue which develops additional tumors and where mucositis is induced after BNCT. We previously reported LAT1 expression in these tumors, in the precancerous and normal pouch tissue. The aim of the present study was to evaluate LAT1 and other molecules of interest in HNC like GLUT1 (a potential transporter for boron compounds and biomarker of aggressiveness), Galectins (GAL1 and GAL3, immunomodulators) and its DNA repair machinery. We also assess if BPA/BNCT affects the expression of these molecules in this oral cancer model. Methods: Tumor-bearing hamsters were divided in [no BPA, 3 animals], [BPA 300 mg/kg euthanized at 3h post injection, 3 animals], [SHAM irradiation: BPA 300 mg/kg euthanized at 1h/4h/24h/10 days post sham irradiation, 3 animals per time point], [BNCT: BPA 300 mg/kg euthanized at 1h/4h/24h/10 days post irradiation, 3 animals per time point]. Tumor bearing and contralateral normal pouches were excised for immunohistochemistry (IHC) and q-PCR for gene expression evaluation. Results: LAT1 IHC and q-PCR results showed that tumor cells are strongly LAT1 positive, with increased LAT1 expression with BPA treatment (p<0.05). LAT1 was also IHC positive in the precancerous and normal pouch tissue. GLUT1 and GAL1 were also upregulated with BPA treatment. Sham and BNCT groups are under study. Conclusions: BPA could be modulating LAT1, GLUT1 and GAL1 in the hamster cheek pouch oral cancer model. LAT1 IHC in the basal epithelium of precancerous and normal tissue was also positive, explaining the accumulation of BPA and consequent mucositis after BNCT. This study provides additional evidence that the hamster oral cancer model is suitable for evaluating BNCT, tumor progression and immunobiomarkers, being useful for the design and development of new BNCT strategies.

We utilized the neutron autoradiography technique to establish a novel approach for detecting B-10 in integrated circuits (ICs). Boron is usually found in the insulating layers of CMOS devices, and its interaction with thermal neutrons is a primary source of ionizing radiation that leads to damage in the chips. Moreover, in order to develop a new technique to detect boron in biological samples for Boron Neutron Capture Therapy (BNCT), it is essential to analyze the presence of boron in CMOS image sensors intended for this purpose. 3 commercial CMOS image sensor models were evaluated through neutron autoradiography in search of boron: OV5647, IMX219, and IMX708. Additionally, thermal neutron irradiations were conducted in order to analyze the events produced by the charged particles resulting from the boron neutron capture reaction and the damage generated by them. A significant presence of boron was found in sensors 0V5647 and IMX219. IMX708 exhibited a minimal presence of boron, making it an attractive candidate for developing the new boron imaging technique for BNCT. The method presented in this work has been proven effective in predicting thermal neutron-induced effects caused by boron in electronic devices and could be applied to any IC, including RAM memories.

Background: Boron neutron capture therapy (BNCT) is a tumor-selective particle radiotherapy that combines preferential boron accumulation in tumors and neutron irradiation. Based on previous studies in tumor-bearing mice, this study evaluated the biodistribution of the sodium salt of cobaltabis(dicarbollide) (Na[3,3′-Co(C2B9H11)2], abbreviated as Na[o-COSAN]) in the hamster cheek pouch oral cancer model and the Na[o-COSAN]/BNCT therapeutic effect on tumors and induced radiotoxicity. The synthesis and comprehensive characterization of 10B-enriched trimethylammonium salt of nido-[7,8-C210B9H12]−o-carborane, along with the cesium and sodium salts of [o-10COSAN] cobaltabis(dicarbollide) are reported here for the first time. Methods: Hamsters bearing tumors were injected with Na[o-COSAN] (7.5 mg B/kg) and euthanized at different time-points after injection (30 min, 2, 3, 5, and 18 h post-administration) to evaluate boron uptake in different tissues/organs. Based on these results, tumor-bearing animals were treated with Na[10B-o-COSAN]/BNCT (7.5 mg B/kg b.w., 3 h), prescribing 5 Gy total in absorbed dose to the precancerous tissue surrounding tumors, i.e., the dose-limiting tissue. Results: Na[o-10COSAN] exhibited no toxicity. Although biodistribution studies employing Na[o-COSAN] have shown low absolute boron concentration in the tumor (approx. 11 ppm), Na[o-10COSAN]/BNCT induced a high and significant therapeutic effect on tumors versus the control group (cancerized, untreated animals). Moreover, only half of the animals exhibited severe mucositis in the precancerous dose-limiting tissue after BNCT, which resolved completely at 21 days after irradiation. Conclusions: Na[o-10COSAN] would be potentially useful to treat head and neck cancer with BNCT.

Boron neutron capture therapy (BNCT) is based on the preferential uptake of 10B compounds by tumors, followed by neutron irradiation. The aim of this study was to assess, in an ectopic colon cancer model, the therapeutic efficacy, radiotoxicity, abscopal effect and systemic immune response associated with (BPA/Borophenylalanine+GB-10/Decahydrodecaborate)-BNCT (Comb-BNCT) alone or in combination with Oligo-Fucoidan (O-Fuco) or Glutamine (GLN), compared to the “standard” BPA-BNCT protocol usually employed in clinical trials. All treatments were carried out at the RA-3 nuclear reactor. Boron biodistribution studies showed therapeutic values above 20 ppm 10B in tumors. At 7 weeks post-treatment, the ratio of tumor volume post-/pre-BNCT was significantly smaller for all BNCT groups vs. SHAM (p < 0.05). The parameter “incidence of tumors that underwent a reduction to ≤50% of initial tumor volume” exhibited values of 62% for Comb-BNCT alone, 82% for Comb-BNCT+GLN, 73% for Comb-BNCT+O-Fuco and only 30% for BPA-BNCT. For BPA-BNCT, the incidence of severe dermatitis was 100%, whereas it was significantly below 70% (p ≤ 0.05) for Comb-BNCT, Comb-BNCT+O-Fuco and Comb-BNCT+GLN. Considering tumors outside the treatment area, 77% of Comb-BNCT animals had a tumor volume lower than 50 mm3 vs. 30% for SHAM (p ≤ 0.005), suggesting an abscopal effect of Comb-BNCT. Inhibition of metastatic spread to lymph nodes was observed in all Comb-BNCT groups. Considering systemic aspects, CD8+ was elevated for Comb-BNCT+GLN vs. SHAM (p ≤ 0.01), and NK was elevated for Comb-BNCT vs. SHAM (p ≤ 0.05). Comb-BNCT improved therapeutic efficacy and reduced radiotoxicity compared to BPA-BNCT and induced an immune response and an abscopal effect.

Boron neutron capture therapy (BNCT) combines preferential tumor uptake of 10B compounds and neutron irradiation. Electroporation induces an increase in the permeability of the cell membrane. We previously demonstrated the optimization of boron biodistribution and microdistribution employing electroporation (EP) and decahydrodecaborate (GB-10) as the boron carrier in a hamster cheek pouch oral cancer model. The aim of the present study was to evaluate if EP could improve tumor control without enhancing the radiotoxicity of BNCT in vivo mediated by GB-10 with EP 10 min after GB-10 administration. Following cancerization, tumor-bearing hamster cheek pouches were treated with GB-10/BNCT or GB-10/BNCT + EP. Irradiations were carried out at the RA-3 Reactor. The tumor response and degree of mucositis in precancerous tissue surrounding tumors were evaluated for one month post-BNCT. The overall tumor response (partial remission (PR) + complete remission (CR)) increased significantly for protocol GB-10/BNCT + EP (92%) vs. GB-10/BNCT (48%). A statistically significant increase in the CR was observed for protocol GB-10/BNCT + EP (46%) vs. GB-10/BNCT (6%). For both protocols, the radiotoxicity (mucositis) was reversible and slight/moderate. Based on these results, we concluded that electroporation improved the therapeutic efficacy of GB-10/BNCT in vivo in the hamster cheek pouch oral cancer model without increasing the radiotoxicity.

Glioblastoma (GBM), as the most central nervous system (CNS) intractable disease, has spoiled millions of lives due to its high mortality. Even though several efforts have been made, the existing treatments have had limited success. In this sense, we studied a lead compound, the boron-rich selective epidermal growth factor receptor (EGFR)-inhibitor hybrid 1, as a potential drug for GBM treatment. For this end, we analyzed the in vitro activity of hybrid 1 in a glioma/primary astrocytes coculture, studying cellular death types triggered by treatment with this compound and its cellular localizations. Additionally, hybrid 1 concentrated boron in glioma cells selectively and more effectively than the boron neutron capture therapy (BNCT)-clinical agent 10B-l-boronophenylalanine and thus displayed a better in vitro-BNCT effect. This encouraged us to analyze hybrid 1 in vivo. Therefore, immunosuppressed mice bearing U87 MG human GBM were treated with both 1 and 1 encapsulated in a modified liposome (recognized by brain–blood barrier peptide transporters), and we observed a potent in vivo per se antitumor activity (tumor size decrease and animal survival increase). These data demonstrate that 1 could be a promising new targeted therapy for GBM.

BACKGROUND:BNCT (Boron Neutron Capture Therapy) is a tumor-selective particle radiotherapy that combines preferential boron accumulation in tumors and neutron irradiation. Although p-boronophenylalanine (BPA) has been clinically used, new boron compounds are needed for the advancement of BNCT. Based on previous studies in colon tumor-bearing mice, in this study, we evaluated MID:BSA (maleimide-functionalized closo-dodecaborate conjugated to bovine serum albumin) biodistribution and MID:BSA/BNCT therapeutic effect on tumors and associated radiotoxicity in the hamster cheek pouch oral cancer model.METHODS:Biodistribution studies were performed at 30 mg B/kg and 15 mg B/kg (12 h and 19 h post-administration). MID:BSA/BNCT (15 mg B/kg, 19 h) was performed at three different absorbed doses to precancerous tissue.RESULTS:MID:BSA 30 mg B/kg protocol induced high BSA toxicity. MID:BSA 15 mg B/kg injected at a slow rate was well-tolerated and reached therapeutically useful boron concentration values in the tumor and tumor/normal tissue ratios. The 19 h protocol exhibited significantly lower boron concentration values in blood. MID:BSA/BNCT exhibited a significant tumor response vs. the control group with no significant radiotoxicity.CONCLUSIONS:MID:BSA/BNCT would be therapeutically useful to treat oral cancer. BSA toxicity is a consideration when injecting a compound conjugated to BSA and depends on the animal model studied.

OBJECTIVE The aim of the present study was to evaluate the local and regional therapeutic efficacy and abscopal effect of BNCT mediated by boronophenyl-alanine, combined with Bacillus Calmette-Guerin (BCG) as an immunotherapy agent in this model. METHODS The local effect of treatment was evaluated in terms of tumor response in the irradiated tumor-bearing right hind flank. Metastatic spread to tumor-draining lymph nodes was analyzed as an indicator of regional effect. The abscopal effect of treatment was assessed as tumor growth inhibition in the contralateral (non-irradiated) left hind flank inoculated with tumor cells 2 weeks post-irradiation. The experimental groups BNCT, BNCT + BCG, BCG, Beam only (BO), BO +BCG, SHAM (tumor-bearing, no treatment, same manipulation) were studied. RESULTS BNCT and BNCT + BCG induced a highly significant local anti-tumor response, whereas BCG alone induced a weak local effect. BCG and BNCT + BCG induced a significant abscopal effect in the contralateral non-irradiated leg. The BNCT + BCG group showed significantly less metastatic spread to tumor-draining lymph nodes vs SHAM and vs BO. CONCLUSION This study suggests that BNCT + BCG-immunotherapy would induce local, regional and abscopal effects in tumor-bearing animals. BNCT would be the main effector of the local anti-tumor effect whereas BCG would be the main effector of the abscopal effect. ADVANCES IN KNOWLEDGE Although the local effect of BNCT has been widely evidenced, this is the first study to show the local, regional and abscopal effects of BNCT combined with immunotherapy, contributing to comprehensive cancer treatment with combined therapies.

One of the driving forces of carcinogenesis in humans is the aberrant activation of receptors; consequently, one of the most promising mechanisms for cancer treatment is receptor inhibition by chemotherapy. Although a variety of cancers are initially susceptible to chemotherapy, they eventually develop multi-drug resistance. Anti-tumor agents overcoming resistance and acting through two or more ways offer greater therapeutic benefits over single-mechanism entities. In this study, we report on a new family of bifunctional compounds that, offering the possibility of dual action (drug + radiotherapy combinations), may result in significant clinical benefits. This new family of compounds combines two fragments: the drug fragment is a lapatinib group, which inhibits the tyrosine kinase receptor activity, and an icosahedral boron cluster used as agents for neutron capture therapy (BNCT). The developed compounds were evaluated in vitro against different tyrosine kinase receptors (TKRs)-expressing tumoral cells, and in vitro–BNCT experiments were performed for two of the most promising hybrids, 19 and 22. We identified hybrid 19 with excellent selectivity to inhibit cell proliferation and ability to induce necrosis/apoptosis of glioblastoma U87 MG cell line. Furthermore, derivative 22, bearing a water-solubility-enhancing moiety, showed moderate inhibition of cell proliferation in both U87 MG and colorectal HT-29 cell lines. Additionally, the HT-29 cells accumulated adequate levels of boron after hybrids 19 and 22 incubations rendering, and after neutron irradiation, higher BNCT-effects than BPA. The attractive profile of developed hybrids makes them interesting agents for combined therapy.

OBJECTIVE(S):The hamster carcinogenesis model recapitulates oral oncogenesis. Dimethylbenz[a]anthracene (DMBA) cancerization induces early severe mucositis, affecting animal's welfare and causing tissue loss and pouch shortening. "Short" pouches cannot be everted for local irradiation for boron neutron capture therapy (BNCT). Our aim was to optimize the DMBA classical cancerization protocol to avoid severe mucositis, without affecting tumor development. We evaluated BNCT in animals cancerized with this novel protocol.MATERIALS AND METHODS:We studied: Classical cancerization protocol (24 applications) and Classical with two interruptions (completed at the end of the cancerization protocol). BNCT mediated by boronophenylalanine (BPA) was performed in both groups.RESULTS:The twice-interrupted group exhibited a significantly lower percentage of animals with severe mucositis versus the non-interrupted group (17% versus 71%) and a significantly higher incidence of long pouches (100% versus 53%). Tumor development and the histologic characteristics of tumor and precancerous tissue were not affected by the interruptions. For both groups, overall tumor response was more than 80%, with a similar incidence of BNCT-induced severe mucositis.CONCLUSION(S):The twice-interrupted protocol reduced severe mucositis during cancerization without affecting tumor development. This favored the animal's welfare and reduced the number of animals to be cancerized for our studies, without affecting BNCT response.

Neutron flux monitoring in research reactors can range from shutdown to full power over 10 to 12 decades. At low power, neutron flux is usually measured with proportional counters in pulse mode. The detector is moved away from high flux zones to avoid pulse saturation until ionization chambers are in range. Campbell mode allows to make a measurement using a single detector able to cover the entire operating range. This work presents an FPGA-based system that implements Campbell mode operation. The system prototype was successfully tested in the CNEA RA-3 research and production reactor at Ezeiza Atomic Center.

Boron neutron capture therapy (BNCT) is a promising cancer binary therapy modality that utilizes the nuclear capture reaction of thermal neutrons by boron-10 resulting in a localized release of high- and low-linear energy transfer (LET) radiation. Electrochemotherapy (ECT) is based on electroporation (EP) that induces opening of pores in cell membranes, allowing the entry of compounds. Because EP is applied locally to a tumor, the compound is incorporated preferentially by tumor cells. Based on the knowledge that the therapeutic success of BNCT depends centrally on the boron content in tumor and normal tissues and that EP has proven to be an excellent facilitator of tumor biodistribution of an anti-tumor agent, the aim of this study was to evaluate if EP can optimize the delivery of boronated compounds. We performed biodistribution studies and qualitative microdistribution analyses of boron employing the boron compound sodium decahydrodecaborate (GB-10) + EP in the hamster cheek pouch oral cancer model. Syrian hamsters with chemically induced exophytic squamous cell carcinomas were used. A typical EP treatment was applied to each tumor, varying the moment of application with respect to the administration of GB-10 (early or late). The results of this study showed a significant increase in the absolute and relative tumor boron concentration and optimization of the qualitative microdistribution of boron by the use of early EP + GB-10 versus GB-10 without EP. This strategy could be a tool to improve the therapeutic efficacy of BNCT/GB-10 in vivo.

Boron neutron capture therapy (BNCT) is a targeted therapy, which consists of preferential accumulation of boron carriers in tumor followed by neutron irradiation. Each oral cancer patient has different risks of developing one or more carcinomas and/or oral mucositis induced after treatment. Our group proposed the hamster oral cancer model to study the efficacy of BNCT and associated mucositis. Translational studies are essential to the advancement of novel boron delivery agents and targeted strategies. Herein, we review our work in the hamster model in which we studied BNCT induced mucositis using three different cancerization protocols, mimicking three different clinical scenarios. The BNCT-induced mucositis increases with the aggressiveness of the carcinogenesis protocol employed, suggesting that the study of different oral cancer patient scenarios would help to develop personalized therapies.

The neutron autoradiography technique using polycarbonate nuclear track detectors (NTD) has been extended to quantify the boron concentration in hard tissues, an application of special interest in Boron Neutron Capture Therapy (BNCT). Chemical and mechanical processing methods to prepare thin tissue sections as required by this technique have been explored. Four different decalcification methods governed by slow and fast kinetics were tested in boron-loaded bones. Due to the significant loss of the boron content, this technique was discarded. On the contrary, mechanical manipulation to obtain bone powder and tissue sections of tens of microns thick proved reproducible and suitable, ensuring a proper conservation of the boron content in the samples. A calibration curve that relates the B-10 concentration of a bone sample and the track density in a Lexan NTD is presented. Bone powder embedded in boric acid solution with known boron concentrations between 0 and 100 ppm was used as a standard material. The samples, contained in slim Lexan cases, were exposed to a neutron fluence of 10(12) cm(-2 )at the thermal column central facility of the RA-3 reactor (Argentina). The revealed tracks in the NTD were counted with an image processing software. The effect of track overlapping was studied and corresponding corrections were implemented in the presented calibration curve. Stochastic simulations of the track densities produced by the products of the B-10 thermal neutron capture reaction for different boron concentrations in bone were performed and compared with the experimental results. The remarkable agreement between the two curves suggested the suitability of the obtained experimental calibration curve. This neutron autoradiography technique was finally applied to determine the boron concentration in pul-verized and compact bone samples coming from a sheep experimental model. The obtained results for both type of samples agreed with boron measurements carried out by ICP-OES within experimental uncertainties. The fact that the histological structure of bone sections remains preserved allows for future boron microdistribution analysis.