Andrea Monti Hughes

“Sequential” Boron Neutron Capture Therapy (BNCT): A Novel Approach to BNCT for the Treatment of Oral Cancer in the Hamster Cheek Pouch Model

In the present study the therapeutic effect and potential toxicity of the novel “Sequential” boron neutron capture therapy (Seq-BNCT) for the treatment of oral cancer was evaluated in the hamster cheek pouch model at the RA-3 Nuclear Reactor. Two groups of animals were treated with “Sequential” BNCT, i.e., BNCT mediated by boronophenylalanine (BPA) followed by BNCT mediated by sodium decahydrodecaborate (GB-10) either 24 h (Seq-24h-BNCT) or 48 h (Seq-48h-BNCT) later. In an additional group of animals, BPA and GB-10 were administered concomitantly [(BPA + GB-10)-BNCT]. The single-application BNCT was to the same total physical tumor dose as the “Sequential” BNCT treatments. At 28 days post-treatment, Seq-24h-BNCT and Seq-48h-BNCT induced, respectively, overall tumor responses of 95 ± 2% and 91 ± 3%, with no statistically significant differences between protocols. Overall response for the single treatment with (BPA + GB-10)-BNCT was 75 ± 5%, significantly lower than for Seq-BNCT. Both Seq-BNCT protocols and (BPA + GB-10)-BNCT induced reversible mucositis in the dose-limiting precancerous tissue around treated tumors, reaching Grade 3/4 mucositis in 47 ± 12% and 60 ± 22% of the animals, respectively. No normal tissue toxicity was associated with tumor response for any of the protocols. “Sequential” BNCT enhanced tumor response without an increase in mucositis in dose-limiting precancerous tissue.

Boron neutron capture therapy (BNCT) for the treatment of liver metastases: biodistribution studies of boron compounds in an experimental model

We previously demonstrated the therapeutic efficacy of different boron neutron capture therapy (BNCT) protocols in an experimental model of oral cancer. BNCT is based on the selective accumulation of 10B carriers in a tumor followed by neutron irradiation. Within the context of exploring the potential therapeutic efficacy of BNCT for the treatment of liver metastases, the aim of the present study was to perform boron biodistribution studies in an experimental model of liver metastases in rats. Different boron compounds and administration conditions were assayed to determine which administration protocols would potentially be therapeutically useful in in vivo BNCT studies at the RA-3 nuclear reactor. A total of 70 BDIX rats were inoculated in the liver with syngeneic colon cancer cells DHD/K12/TRb to induce the development of subcapsular tumor nodules. Fourteen days post-inoculation, the animals were used for biodistribution studies. We evaluated a total of 11 administration protocols for the boron compounds boronophenylalanine (BPA) and GB-10 (Na210B10H10), alone or combined at different dose levels and employing different administration routes. Tumor, normal tissue, and blood samples were processed for boron measurement by atomic emission spectroscopy. Six protocols proved potentially useful for BNCT studies in terms of absolute boron concentration in tumor and preferential uptake of boron by tumor tissue. Boron concentration values in tumor and normal tissues in the liver metastases model show it would be feasible to reach therapeutic BNCT doses in tumor without exceeding radiotolerance in normal tissue at the thermal neutron facility at RA-3.

Therapeutic efficacy of boron neutron capture therapy mediated by boron-rich liposomes for oral cancer in the hamster cheek pouch model

Significance Boron neutron capture therapy (BNCT) for cancer is based on the selective uptake of 10B target compounds by tumor cells followed by neutron irradiation. The capture reaction between 10B atoms and neutrons gives rise to short-range particles, which are highly effective in producing cell damage. Thus, BNCT is designed to damage tumor cells and preserve healthy cells. The boron carrier used is pivotal to the success of BNCT. The present study describes the therapeutic success of BNCT mediated by MAC-TAC liposomes, K[nido-7-CH3(CH2)15-7,8-C2B9H11] (MAC) in the bilayer membrane and encapsulating the hydrophilic species Na3[ae-B20H17NH3] (TAC) in the aqueous core, using the hamster cheek pouch oral cancer model. A sustained tumor response of 70–88% was associated with only mild mucositis in dose-limiting precancerous tissue. The application of boron neutron capture therapy (BNCT) mediated by liposomes containing 10B-enriched polyhedral borane and carborane derivatives for the treatment of head and neck cancer in the hamster cheek pouch oral cancer model is presented. These liposomes are composed of an equimolar ratio of cholesterol and 1,2-distearoyl-sn-glycero-3-phosphocholine, incorporating K[nido-7-CH3(CH2)15-7,8-C2B9H11] (MAC) in the bilayer membrane while encapsulating the hydrophilic species Na3[ae-B20H17NH3] (TAC) in the aqueous core. Unilamellar liposomes with a mean diameter of 83 nm were administered i.v. in hamsters. After 48 h, the boron concentration in tumors was 67 ± 16 ppm whereas the precancerous tissue contained 11 ± 6 ppm, and the tumor/normal pouch tissue boron concentration ratio was 10:1. Neutron irradiation giving a 5-Gy dose to precancerous tissue (corresponding to 21 Gy in tumor) resulted in an overall tumor response (OR) of 70% after a 4-wk posttreatment period. In contrast, the beam-only protocol gave an OR rate of only 28%. Once-repeated BNCT treatment with readministration of liposomes at an interval of 4, 6, or 8 wk resulted in OR rates of 70–88%, of which the complete response ranged from 37% to 52%. Because of the good therapeutic outcome, it was possible to extend the follow-up of BNCT treatment groups to 16 wk after the first treatment. No radiotoxicity to normal tissue was observed. A salient advantage of these liposomes was that only mild mucositis was observed in dose-limiting precancerous tissue with a sustained tumor response of 70–88%.

Tumor Blood Vessel “Normalization” Improves the Therapeutic Efficacy of Boron Neutron Capture Therapy (BNCT) in Experimental Oral Cancer

We previously demonstrated the efficacy of BNCT mediated by boronophenylalanine (BPA) to treat tumors in a hamster cheek pouch model of oral cancer with no normal tissue radiotoxicity and moderate, albeit reversible, mucositis in precancerous tissue around treated tumors. It is known that boron targeting of the largest possible proportion of tumor cells contributes to the success of BNCT and that tumor blood vessel normalization improves drug delivery to the tumor. Within this context, the aim of the present study was to evaluate the effect of blood vessel normalization on the therapeutic efficacy and potential radiotoxicity of BNCT in the hamster cheek pouch model of oral cancer. Blood vessel normalization was induced by two doses of thalidomide in tumor-bearing hamsters on 2 consecutive days. All studies in thalidomide-treated animals were performed 48 h after the first dose of thalidomide, previously established as the window of normalization. Biodistribution studies were performed with BPA at a dose of 15.5 mg 10B/kg in thalidomide-treated (Th+) and untreated (Th–) tumor-bearing hamsters. The effect of blood vessel normalization prior to BPA administration on the efficacy of BNCT was assessed in in vivo BNCT studies at the RA-3 Nuclear Reactor in tumor-bearing hamsters. Group I was treated with BPA-BNCT after treatment with thalidomide (Th+ BPA-BNCT). Group II was treated with BPA-BNCT alone (Th– BPA-BNCT). Group III was treated with the beam only after treatment with thalidomide (Th+ BO), and Group IV was treated with the beam only (Th– BO). Groups I and II were given the same dose of BPA (15.5 mg 10B/kg), and all groups (I–IV) were exposed to the same neutron fluence. Two additional groups were treated with the beam only at a higher dose to exacerbate mucositis in precancerous tissue and to explore the potential direct protective effect of thalidomide on radiation-induced mucositis in a scenario of more severe toxicity, i.e. Group V (Th+ hdBO) and Group VI (Th– hdBO). The animals were followed for 28 days. Biodistribution studies revealed no statistically significant differences in gross boron content between Th+ and Th– animals. Overall tumor control (complete response + partial response) at 28 days post-treatment was significantly higher for Group I (Th+ BPA-BNCT) than for Group II (Th– BPA-BNCT): 84 ± 3% compared to 67 ± 5%. Pretreatment with thalidomide did not induce statistically significant changes in overall tumor control induced by the beam only, i.e. 15 ± 5% in Group III (Th+ BO) and 18 ± 5% in Group IV (Th– BO), or in overall tumor control induced by the high-dose beam only, i.e. 60 ± 7% in Group V (Th+ hdBO) and 47 ± 10% in Group VI (Th– hdBO). BPA-BNCT alone (Group II) induced mucositis in precancerous tissue that reached Grades 3–4 in 80% of the animals, whereas pretreatment with thalidomide (Group I) prevented mucositis Grades 3 and 4 completely. Beam-only Group III (Th+ BO) exhibited only Grade 1 mucositis in precancerous tissue, whereas 17% of the animals in beam-only Group IV (Th– BO) reached Grade 2 mucositis. High-dose beam-only group V (Th+ hdBO) exhibited only Grade 2 mucositis, whereas high-dose beam-only group VI (Th– hdBO) reached Grade 3 mucositis in 83% of the animals. In all cases mucositis in precancerous tissue was reversible. No normal tissue radiotoxicity was observed with any of the protocols. Pretreatment with thalidomide enhanced the therapeutic efficacy of BNCT and reduced precancerous tissue toxicity.

Development of a model of tissue with potentially malignant disorders (PMD) in the hamster cheek pouch to explore the long-term potential therapeutic and/or toxic effects of different therapeutic modalities

OBJECTIVE Given that locoregional recurrences developing from a tissue with potentially malignant disorders (PMD) in oral mucosa are a frequent cause of therapeutic failure, and that tissue with PMD is dose-limiting, the aim of the present study was to develop a model of tissue with PMD to evaluate the long-term therapeutic/toxic effects of different therapeutic modalities. MATERIALS AND METHODS We evaluated 5 carcinogenesis protocols based on topical application of the carcinogen dimethyl-1,2-benzanthracene in the hamster cheek pouch, twice a week for 4, 6, 7, and 8 weeks and the classical 3 times a week for 12 weeks. RESULTS Long-term follow-up (8 months after protocol completion) was only possible with the 4- and 6-week carcinogenesis protocols. Tumour development increased progressively with time and aggressiveness of the carcinogenesis protocols. The time at which tumours developed in > or =90% of the animals was at protocol completion (T0) for the 12-week protocol, 1 month post-T0 for the 8-week protocol, 3 months post-T0 for the 7-week protocol and 4 months post-T0 for the 6-week protocol. <40% of the animals in the 4-week protocol developed tumours within the 8 months follow-up period. DNA synthesis rose as a function of time and protocol aggressiveness. CONCLUSIONS The 6-week carcinogenesis protocol was selected for long-term studies of different therapeutic modalities in tissue with PMD because it permitted long-term follow-up and guaranteed tumour development in > or =90% of the animals.

Boron Neutron Capture Therapy (BNCT) in an oral precancer model: Therapeutic benefits and potential toxicity of a double application of BNCT with a six-week interval

Given the clinical relevance of locoregional recurrences in head and neck cancer, we developed a novel experimental model of premalignant tissue in the hamster cheek pouch for long-term studies and demonstrated the partial inhibitory effect of a single application of Boron Neutron Capture Therapy (BNCT) on tumor development from premalignant tissue. The aim of the present study was to evaluate the effect of a double application of BNCT with a 6 week interval in terms of inhibitory effect on tumor development, toxicity and DNA synthesis. We performed a double application, 6 weeks apart, of (1) BNCT mediated by boronophenylalanine (BPA-BNCT); (2) BNCT mediated by the combined application of decahydrodecaborate (GB-10) and BPA [(GB-10+BPA)-BNCT] or (3) beam-only, at RA-3 nuclear reactor and followed the animals for 8 months. The control group was cancerized and sham-irradiated. BPA-BNCT, (GB-10+BPA)-BNCT and beam-only induced a reduction in tumor development from premalignant tissue that persisted until 8, 3, and 2 months respectively. An early maximum inhibition of 100% was observed for all 3 protocols. No normal tissue radiotoxicity was detected. Reversible mucositis was observed in premalignant tissue, peaking at 1 week and resolving by the third week after each irradiation. Mucositis after the second application was not exacerbated by the first application. DNA synthesis was significantly reduced in premalignant tissue 8 months post-BNCT. A double application of BPA-BNCT and (GB-10+BPA)-BNCT, 6 weeks apart, could be used therapeutically at no additional cost in terms of radiotoxicity in normal and dose-limiting tissues.

Assessing advantages of sequential boron neutron capture therapy (BNCT) in an oral cancer model with normalized blood vessels

Abstract Background. We previously demonstrated the therapeutic success of sequential boron neutron capture therapy (Seq-BNCT) in the hamster cheek pouch oral cancer model. It consists of BPA-BNCT followed by GB-10-BNCT 24 or 48 hours later. Additionally, we proved that tumor blood vessel normalization with thalidomide prior to BPA-BNCT improves tumor control. The aim of the present study was to evaluate the therapeutic efficacy and explore potential boron microdistribution changes in Seq-BNCT preceded by tumor blood vessel normalization. Material and Methods. Tumor bearing animals were treated with thalidomide for tumor blood vessel normalization, followed by Seq-BNCT (Th+ Seq-BNCT) or Seq-Beam Only (Th+ Seq-BO) in the window of normalization. Boron microdistribution was assessed by neutron autoradiography. Results. Th+ Seq-BNCT induced overall tumor response of 100%, with 87 (4)% complete tumor response. No cases of severe mucositis in dose-limiting precancerous tissue were observed. Differences in boron homogeneity between tumors pre-treated and not pre-treated with thalidomide were observed. Conclusion. Th+ Seq-BNCT achieved, for the first time, response in all treated tumors. Increased homogeneity in tumor boron microdistribution is associated to an improvement in tumor control.

Dynamic infrared imaging for biological and medical applications in Boron neutron capture therapy

Boron Neutron Capture Therapy (BNCT) is a treatment modality, currently focused on the treatment of cancer, which involves a tumor selective 10B compound and a specially tuned neutron beam to produce a lethal nuclear reaction. BNCT kills target cells with microscopic selectivity while sparing normal tissues from potentially lethal doses of radiation. In the context of the Argentine clinical and research BNCT projects at the National Atomic Energy Commission and in a strong collaboration with INVAP SE, we successfully implemented Dynamic Infrared Imaging (DIRI) in the clinical setting for the observation of cutaneous melanoma patients and included DIRI as a non invasive methodology in several research protocols involving small animals. We were able to characterize melanoma lesions in terms of temperature and temperature rate-of-recovery after applying a mild cold thermal stress, distinguishing melanoma from other skin pigmented lesions. We observed a spatial and temporal correlation between skin acute reactions after irradiation, the temperature pattern and the dose distribution. We studied temperature distribution as a function of tumor growth in mouse xenografts, observing a significant correlation between tumor temperature and drug uptake; we investigated temperature evolution in the limbs of Wistar rats for a protocol of induced rheumatoid arthritis (RA), DIRI being especially sensitive to RA induction even before the development of clinical signs and studied surface characteristics of tumors, precancerous and normal tissues in a model of oral cancer in the hamster cheek pouch.

Non-invasive characterization of normal and pathological tissues through dynamic infrared imaging in the hamster cheek pouch oral cancer model

Biomedical infrared thermography, a non-invasive and functional imaging method, provides information on the normal and abnormal status and response of tissues in terms of spatial and temporal variations in body infrared radiance. It is especially attractive in cancer research due to the hypervascular and hypermetabolic activity of solid tumors. Moreover, healthy tissues like skin or mucosa exposed to radiation can be examined since inflammation, changes in water content, exudation, desquamation, erosion and necrosis, between others, are factors that modify their thermal properties. In this work we performed Dynamic Infrared Imaging (DIRI) to contribute to the understanding and evaluation of normal tissue, tumor and precancerous tissue response and radiotoxicity in an in vivo model, the hamster cheek pouch, exposed to Boron Neutron Capture Therapy. In this study, we particularly focused on the observation of temperature changes under forced transient conditions associated with mass moisture transfer in the tissue-air interface, in each tissue with or without treatment. We proposed a simple mathematical procedure that considerers the heat transfer from tissue to ambient through convection and evaporation to model the transient (exponential decay o recover) thermal study. The data was fitted to determined the characteristic decay and recovery time constants of the temperature as a function of time. Also this model allowed to explore the mass flux of moisture, as a degree of evaporation occurring on the tissue surface. Tissue thermal responses under provocation tests could be used as a non-invasive method to characterize tissue physiology.

Boron biodistribution for BNCT in the hamster cheek pouch oral cancer model: Combined administration of BSH and BPA

Sodium mercaptoundecahydro-closo-dodecaborate (BSH) is being investigated clinically for BNCT. We examined the biodistribution of BSH and BPA administered jointly in different proportions in the hamster cheek pouch oral cancer model. The 3 assayed protocols were non-toxic, and showed preferential tumor boron uptake versus precancerous and normal tissue and therapeutic tumor boron concentration values (70-85ppm). All 3 protocols warrant assessment in BNCT studies to contribute to the knowledge of (BSH+BPA)-BNCT radiobiology for head and neck cancer and optimize therapeutic efficacy.