Clinical Viability of Boron Neutron Capture Therapy for Personalized Radiation Treatment

Citation: Skwierawska, D.; López-Valverde, J.A.; Balcerzyk, M.; Leal, A. Clinical Viability of Boron Neutron Capture Therapy for Personalized Radiation Treatment. Cancers 2022,14, 2865. https:// doi.org/10.3390/cancers14122865 Academic Editor: Denise Bernhardt Received: 30 April 2022 Accepted: 8 June 2022 Published: 10 June 2022 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). cancers Review Clinical Viability of Boron Neutron Capture Therapy for Personalized Radiation Treatment Dominika Skwierawska 1,†,‡ , JoséAntonio López-Valverde 2,3,† , Marcin Balcerzyk 2,4,* and Antonio Leal 2,3,* 1Department of Biomedical Engineering, Faculty of Electronics, Telecommunications, and Informatics, Gda´nsk University of Technology, 80-233 Gda´nsk, Poland; [email protected] 2Departamento de Fisiología Médica y Biofísica, Facultad de Medicina, Universidad de Sevilla, 41009 Seville, Spain; [email protected] 3Instituto de Biomedicina de Sevilla, IBiS, 41013 Seville, Spain 4Unidad Ciclotrón, Centro Nacional de Aceleradores, Universidad de Sevilla-CSIC-Junta de Andalucía, 41093 Seville, Spain *Correspondence: [email protected] (M.B.); [email protected] (A.L.) † These authors contributed equally to this work. ‡ Current address: MERK Sp. z o.o., Mikołaja Reja 3, 80-404 Gda´nsk, Poland. Simple Summary: Usually, for dose planning in radiotherapy, the tumor is delimited as a volume on the image of the patient together with other clinical considerations based on populational evidence. However, the same prescription dose can provide different results, depending on the patient. Unfortunately, the biological aspects of the tumor are hardly considered in dose planning. Boron Neutron Capture Radiotherapy enables targeted treatment by incorporating boron-10 at the cellular level and irradiating with neutrons of a certain energy so that they produce nuclear reactions locally and almost exclusively damage the tumor cell. This technique is not new, but modern neutron generators and more efficient boron carriers have reactivated the clinical interest of this technique in the pursuit of more precise treatments. In this work, we review the latest technological facilities and future possibilities for the clinical implementation of BNCT and for turning it into a personalized therapy. Abstract: Boron Neutron Capture Therapy (BNCT) is a promising binary disease-targeted therapy, as neutrons preferentially kill cells labeled with boron ( 10 B), which makes it a precision medicine treatment modality that provides a therapeutic effect exclusively on patient-specific tumor spread. Contrary to what is usual in radiotherapy, BNCT proposes cell-tailored treatment planning rather than to the tumor mass. The success of BNCT depends mainly on the sufficient spatial biodistribution of 10 B located around or within neoplastic cells to produce a high-dose gradient between the tumor and healthy tissue. However, it is not yet possible to precisely determine the concentration of 10 B in a specific tissue in real-time using non-invasive methods. Critical issues remain to be resolved if BNCT is to become a valuable, minimally invasive, and efficient treatment. In addition, functional imaging technologies, such as PET, can be applied to determine biological information that can be used for the combined-modality radiotherapy protocol for each specific patient. Regardless, not only imaging methods but also proteomics and gene expression methods will facilitate BNCT becoming a modality of personalized medicine. This work provides an overview of the fundamental principles, recent advances, and future directions of BNCT as cell-targeted cancer therapy for personalized radiation treatment. Keywords: BNCT; targeted therapy; biological dosimetry; boron imaging; personalized oncology; personalized medicine 1. Introduction Cancer, a multicellular and multigenic disease, is one of the leading causes of death in the world. It is the first/second cause in 112 out of 183 countries and the third/fourth in Cancers 2022,14, 2865. https://doi.org/10.3390/cancers14122865 https://www.mdpi.com/journal/cancers Cancers 2022,14, 2865 2 of 26 23 countries, according to estimated data from the World Health Organization (WHO) in 2019. In 2020, 19.3 million new cases and 10 million cancer deaths were recorded according to GLOBOCAN estimates of 36 cancers in 185 countries [ 1 ]. Given that the annual incidence continues to increase, clinical management of cancer remains a significant challenge. Cancer can arise from all organs and different cell types with a multifactorial etiology. In general, cancer cells exhibit inherent phenotypical characteristics, known as hallmarks of cancer. Hanahan and Weinberg [ 2 ] originally suggested six alterations in cell physiology that collectively dictate malignant growth: environmental independence for growth, evasion of apoptosis (programmed cell death), limitless proliferative potential, sustained angiogenesis, tissue invasion, and metastasis to other parts of the body. In a more recent update, they also included deregulated metabolism and immune system evasion as additional hallmarks, as well as two characteristics that allow the acquisition of all hallmarks: genome instability and inflammation [3]. In countries with a high gross domestic product, radiotherapy (RT) is used in more than 50% of patients to treat the disease in a local stage or to control and alleviate symptoms of irrecoverable cases, depending on the stage of cancer [ 1 ]. Currently, estimates of the global demand for RT by cancer patients indicate that, for 87% of new cases of breast cancer patients, RT is an important part of therapy according to clinical guidelines and evidence-based medicine. In the case of surgery in the early stages of breast cancer and systemic therapies in metastatic cases, RT is applied as an adjunctive therapy [ 4 ]. In several types of tumors, such as head and neck cancers, skin cancers, cervical cancers, brain tumors, and others, RT is considered the alternative curative treatment option or even the basis for definitive curative treatment. Additionally, in many locations of the disease, RT is usually required as a prophylactic agent after surgery. In locations such as the lungs, the precision of the radiation beams allows the use of RT under ablative conditions when surgery is not applicable. This is the case of Stereotactic Ablative Radiotherapy (SABR), where vascular endothelial injury and immune activation are new radiobiological aspects that must be added to the 5 R’s of radiobiology (reoxygenation, repair, radiosensitivity, redistribution, and repopulation) to explain the ablation effect. These data indicate that the role of RT has increased in prominence compared to surgery in the treatment of localized solid tumors, the most widespread expression of cancer. Among other reasons, these achievements are the result of several innovative therapeutic methods and technological improvements, such as the implementation of devices to shape and adapt the irradiation beam to the tumor volume while safeguarding organs at risk, or the successful implementation of advanced imaging procedures used in planning and treatment [ 5 , 6 ]. RT aims to deliver the optimal dose to tumor volume while preserving normal tissues, but these volumes are considered at the macroscopic level in treatment planning following an evidence-based population medicine approach through the image processing for segmentation of the target volume in the image data of patients usually provided by CT devices [ 7 ]. Unfortunately, patients often vary between tumor responses to RT due to differences in tumor type and other specific genetic factors not yet considered in treatment planning. Beyond technological or clinical constraints, the efficacy of RT is strongly limited by the different biological characteristics of the tumor. Personalized medicine is usually related to genetic or other biomarker information to make treatment decisions for individually considered patients. Under this definition, RT cannot play a leading role in personalized cancer treatment as it has been played so far by chemotherapy and immunotherapy. However, efficient synergies between methodologies with a basic scientific profile and other more technical ones that have revolutionized RT could be the key to the real clinical implementation of personalized cancer therapy. In this sense, Boron Neutron Capture Therapy (BNCT) shows interesting possibilities insofar as it provides cellular targeting of radiation energy transfer and so could make more efficient previous genetic and molecular information about each patient. Cancers 2022,14, 2865 3 of 26 BNCT is a binary treatment method based on the combination of two agents, 10 B and epithermal neutrons, that exploits the high linear energy transfer (LET) characteristics of the mixture of fractionation components. Boron can be selectively localized in tumor cells. Thus, BNCT is a promising disease-targeted therapy, as neutrons preferentially kill cells labeled with 10 B, making it a treatment modality of precision medicine. The overall BNCT procedure, indicating the principal steps from diagnosis confirmation to post-treatment monitoring, is shown in Figure 1. Cancers 2022, 14, x 3 of 26 targeting of radiation energy transfer and so could make more efficient previous genetic and molecular information about each patient. BNCT is a binary treatment method based on the combination of two agents, 10B and epithermal neutrons, that exploits the high linear energy transfer (LET) characteristics of the mixture of fractionation components. Boron can be selectively localized in tumor cells. Thus, BNCT is a promising disease-targeted therapy, as neutrons preferentially kill cells labeled with 10B, making it a treatment modality of precision medicine. The overall BNCT procedure, indicating the principal steps from diagnosis confirmation to post-treatment monitoring, is shown in Figure 1. Figure 1. Standard BNCT procedure. From prior medical examination and diagnosis confirmation with the use of [18F]FBPA PET, through Boron Neutron Capture therapy, to post-therapy monitoring and evaluation. Two types of neutron beam are commonly used in BNCT: thermal beams (~0.0254 eV) and epithermal beams (0.5 eV to 40 keV). For clinical purposes, the most useful are epithermal neutrons because, while entering the tissue, they create a radiation field with maximum thermal flux at a depth of 2–3 cm, which then drops exponentially. In turn, when a thermal beam enters the tissue, the thermal flux—which is created as a result— falls exponentially from the surface. A fundamental principle of BNCT method is 10B(n,α)7Li nuclear reaction, which occurs when the stable isotope 10B, which is administered preferentially to tumor cells, is subsequently irradiated with an external epithermal neutron beam to produce an α-particle (4He) and a 7Li nucleus. A schematic representation of this reaction is presented in Figure 2. Released α-particles (~1.47 MeV) and the 7Li nuclei (~0.84 MeV) have a high LET ~175 keV·μm−1 [8–10]. About 94% of the time, the recoiling 7Li ion is produced in an excited state and emits a low LET 477 keV gamma-ray during deexcitation. In the remaining 6% of events, 7Li is emitted with no gamma-ray emission in the ground state. Figure 1. Standard BNCT procedure. From prior medical examination and diagnosis confirmation with the use of [ 18 F]FBPA PET, through Boron Neutron Capture therapy, to post-therapy monitoring and evaluation. Two types of neutron beam are commonly used in BNCT: thermal beams (~0.0254 eV) and epithermal beams (0.5 eV to 40 keV). For clinical purposes, the most useful are epithermal neutrons because, while entering the tissue, they create a radiation field with maximum thermal flux at a depth of 2–3 cm, which then drops exponentially. In turn, when a thermal beam enters the tissue, the thermal flux—which is created as a result—falls exponentially from the surface. A fundamental principle of BNCT method is 10 B(n, α ) 7 Li nuclear reaction, which occurs when the stable isotope 10 B, which is administered preferentially to tumor cells, is subsequently irradiated with an external epithermal neutron beam to produce an α - particle ( 4 He) and a 7 Li nucleus. A schematic representation of this reaction is presented in Figure 2. Released α -particles (~1.47 MeV) and the 7 Li nuclei (~0.84 MeV) have a high LET ~175 keV·µm−1[8–10]. About 94% of the time, the recoiling 7 Li ion is produced in an excited state and emits a low LET 477 keV gamma-ray during deexcitation. In the remaining 6% of events, 7Li is emitted with no gamma-ray emission in the ground state. Unfortunately, after many years of research led by scientists and specialists, they are still struggling with some critical issues of BNCT. Firstly, generating a therapeutic beam with an optimal energy spectrum that can deliver the neutrons to the correct location while minimizing the dose delivered to healthy tissue. Second, finding non-toxic 10 B delivery agents: the boron carrier compound is one of the fundamental aspects of BNCT and should bring significantly more 10 B to the cancerous cells than to the healthy tissue or, ideally, Cancers 2022,14, 2865 4 of 26 only into the cancerous cells. It should also meet requirements, such as water solubility, chemical stability, and preservation of constant high concentration during the treatment procedure [ 11 ]. Because the boron concentration level directly affects the yield of alpha and Li particles generated and hence the dose to the tumor and other tissue, it is also essential to image the boron distribution when considering BNCT. Several modalities are widely used to assess the boron dose delivered to the residual tumor volume, and they can provide information on the distribution of 10 B at the microscopic level [ 12 , 13 ]. To improve molecular imaging, several other approaches have been proposed [ 14 , 15 ]. Another critical issue is developing treatment planning programs and systems to calculate the dose, predict particle fluxes, and expect the incidence angles in patients to achieve an adequate relative dose distribution. If the reactor generates the BNCT treatment beams, the treatment planner must determine the dose induced by neutrons and gamma photons. In this case, most of the gamma photons that occur in the beam originate from the reactor core. Fortunately, development research on compact, in-hospital accelerator-based neutron sources, ready for installation in hospital environments, has been ongoing for many years in several countries. It allowed for more widespread use of the BNCT technique and removed the contamination from high energy gammas from the reactor core, which was one of the reasons for the failure of BNCT in the 1950s. Cancers 2022, 14, x 4 of 26 Figure 2. 10B (n, α)7Li reaction. Unfortunately, after many years of research led by scientists and specialists, they are still struggling with some critical issues of BNCT. Firstly, generating a therapeutic beam with an optimal energy spectrum that can deliver the neutrons to the correct location while minimizing the dose delivered to healthy tissue. Second, finding non-toxic 10B delivery agents: the boron carrier compound is one of the fundamental aspects of BNCT and should bring significantly more 10B to the cancerous cells than to the healthy tissue or, ideally, only into the cancerous cells. It should also meet requirements, such as water solubility, chemical stability, and preservation of constant high concentration during the treatment procedure [11]. Because the boron concentration level directly affects the yield of alpha and Li particles generated and hence the dose to the tumor and other tissue, it is also essential to image the boron distribution when considering BNCT. Several modalities are widely used to assess the boron dose delivered to the residual tumor volume, and they can provide information on the distribution of 10B at the microscopic level [12,13]. To improve molecular imaging, several other approaches have been proposed [14,15]. Another critical issue is developing treatment planning programs and systems to calculate the dose, predict particle fluxes, and expect the incidence angles in patients to achieve an adequate relative dose distribution. If the reactor generates the BNCT treatment beams, the treatment planner must determine the dose induced by neutrons and gamma photons. In this case, most of the gamma photons that occur in the beam originate from the reactor core. Fortunately, development research on compact, in-hospital accelerator-based neutron sources, ready for installation in hospital environments, has been ongoing for many years in several countries. It allowed for more widespread use of the BNCT technique and removed the contamination from high energy gammas from the reactor core, which was one of the reasons for the failure of BNCT in the 1950s. Numerous reasons define why the cell may die. They relate to the cell cycle phase in which irradiation occurs, the cell type, the radiation dose, and physiological conditions, such as oxygen supply. However, what drives cancer cells during the switch from a repair program to cell death and what drives the cancerous cell to choose a specific pathway of death? The specific mechanism of cell death and the mechanism of repair after BNCT are not sufficiently known. In this case, the evaluation of early and late markers of cellular responses after the introduction of BNCT should be considered. They are crucial for the further development of BNCT. BNCT could be treated as a personalized treatment, as 10B uptake depends on factors such as the characteristics of the individual tumor cell, the pharmacokinetics of boron drugs, and its subcellular distribution. Moreover, the optimal moment of boron-carrying drug concentration ratio in tumor cells vs. healthy ones, which varies from patient to patient, must be achieved. This review highlights the issues described above and evaluates them in future directions and the further development of BNCT as an effective personalized radiation therapy. The answers to the questions presented and others that will undoubtedly arise in the further development of research on cancer radiation therapy, and the progression in solving the issues presented, even in part, will eventually lead to continued improvement in BNCT-based cancer treatment. Figure 2. 10B (n,α)7Li reaction. Numerous reasons define why the cell may die. They relate to the cell cycle phase in which irradiation occurs, the cell type, the radiation dose, and physiological conditions, such as oxygen supply. However, what drives cancer cells during the switch from a repair program to cell death and what drives the cancerous cell to choose a specific pathway of death? The specific mechanism of cell death and the mechanism of repair after BNCT are not sufficiently known. In this case, the evaluation of early and late markers of cellular responses after the introduction of BNCT should be considered. They are crucial for the further development of BNCT. BNCT could be treated as a personalized treatment, as 10 B uptake depends on factors such as the characteristics of the individual tumor cell, the pharmacokinetics of boron drugs, and its subcellular distribution. Moreover, the optimal moment of boron-carrying drug concentration ratio in tumor cells vs. healthy ones, which varies from patient to patient, must be achieved. This review highlights the issues described above and evaluates them in future directions and the further development of BNCT as an effective personalized radiation therapy. The answers to the questions presented and others that will undoubtedly arise in the further development of research on cancer radiation therapy, and the progression in solving the issues presented, even in part, will eventually lead to continued improvement in BNCT-based cancer treatment. 2. Fundamental Aspects of BNCT BNCT is primarily a biochemically, rather than physically targeted type of radiation therapy, so fundamental aspects are first introduced in this section, such as the characteristics of BNCT as a targeted and binary therapy, the possible mechanisms and types of BNCT-induced cell death that occur during and after irradiation, and to pay Cancers 2022,14, 2865 5 of 26 attention to the necessary biological dosimetry to understand and evaluate the increased radio-sensitization effect. 2.1. BNCT as Targeted Radiotherapy BNCT could enable highly individualized tumor treatment in the sense that the therapeutic effect can be limited to the specific tumor spread of the patient. In BNCT, this will only be possible when a compound accumulates 10 B only in all tumor cells with a sufficiently high concentration in relation to healthy cells. Nuclear localization is preferred to maximize DNA damage. Therefore, boron delivery agents are one of the essential aspects of BNCT. 10 B should be retained in the tumor, at least for the duration of neutron irradiation, which can take up to an hour. However, the way to concentrate 10 B in sufficient amounts and preferentially in cancer cells is currently the main limitation of the effectiveness of BNCT. The first step is the separation of 10 B (20% abundance) from 11 B (80%), which is mastered but expensive process. Many compounds have been developed to date, but currently only two boron agents are widely used as boron carriers: sodium borocaptate (Na 2 B 12 H 11 SH [[ 10 B]BSH]) and [ 10 B]4-borono-L-phenylalanine (BPA), two drugs containing low molecular weight boron. BSH used in BNCT consists of 12 10 B atoms and is used mainly to treat malignant gliomas. BSH is not delivered to the normal brain through the intact blood–brain barrier (BBB), and it is difficult to selectively internalize in tumor cells because of its high hydrophilicity. Its concentration in the target is related to the concentration of the agent in the blood and the vascularization of the neoplasm [ 16 ]. BSH has a passive diffuse accumulation mechanism. In malignant cells in the brain, it accumulates only in the tumor region where the blood–brain barrier is disrupted [17]. On the other hand, BPA is a derivative of phenylalanine and is actively transported into tumor cells, mainly through the L-type amino acid transporter 1 (LAT 1) [ 18 ]. The BPA logP is negative ( − 3.65), and this indicates that it will not pass the BBB passively because only small molecules with logP in the range of 1.5–2.7 can cross the BBB with passive (diffusive) transport [ 19 , 20 ]. Therefore, the only way for BPA to cross the BBB is to sneak it through some transporter, such as an L-type amino acid. BPA has been reported to accumulate specifically in tumor cells due to its structural similarities to tyrosine [ 5 ]. As L-type amino acid transporters are involved in some important human diseases and are overexpressed in human tumors, they improve targeted delivery to the brain and cancer cells. LAT 1 is also present in the blood–brain barrier (BBB), the blood–retina barrier, the cerebral cortex, testes, placenta, and bone marrow. Injection of [ 10 B]BPA for intravenous administration in BNCT is prepared as the [ 10 B]BPA-fructose complex [ 21 ]. The reason for labeling these compounds with positron emitters is to accurately determine the boron distribution and concentration in the tumor and surrounding tissue using PET, as will be discussed in Section 3. BPA was approved in Japan as a commercial drug with social security reimbursement and has been available on the market since 20 May 2020 under the name Borofalan (10B) [22]. Three generations of boron compounds can be distinguished: (I) borax, boric acid, and its derivatives used in the first clinical trials [ 23 , 24 ]; (II) boron-modified amino acids, including boron carriers, such as BPA and BSH; and (III) a third generation of boron agents, which has attracted the attention of scientists over the past two decades. They focus on using biochemical pathways to accumulate boronated analogs in subcellular structures. These new BNCT agents include small molecules containing boron, such as peptides [ 25 , 26 ], antibodybased delivery systems [ 27 ], boron compound conjugates [ 28 – 30 ], boron-dispersed nanoparticles [ 31 – 34 ] (nanomaterial-based delivery systems), and others currently under evaluation. Targeted boron delivery agents combine boron-containing agents with tumor-targeting molecules (e.g., nucleosides, porphyrins, peptides, proteins, or antibodies). Boron-delivery nanomaterials can transport various boron-containing compounds into tumor cells by taking advantage of the enhanced permeability and retention effect that affects nanomaterials and the active targeting effects mediated by tumor-targeted ligands grafted on the surface Cancers 2022,14, 2865 6 of 26 of the materials. The following boron-delivery nanomaterials can be distinguished: dendrimers, liposomes, polymeric nanoparticles, boron nitride, carbon nanotubes, mesoporous silica nanoparticles, ferromagnetic and paramagnetic nanoparticles—some of them used for MRI imaging—, gold nanoparticles, and BPO 4 nanoparticles. There are many promising routes in drug delivery systems, and there is still a pressing need to develop new boron delivery agents, but without adequate research and clinical trials, it is difficult to determine which is the most feasible [35,36]. There are some obstacles to the exploitation of the full potential of the cancer-specific selectivity of BNCT, including the suboptimal boron delivery strategies currently used. However, various supportive targeted measures are under development, for example, delivery agents targeting the glucose transporter GLUT1 [37]. In any case, BNCT workflow would allow for a full personalization of the treatment, from the drug delivery treatment to imaging, and considering high throughput techniques, as it is discussed later in the “BNCT and personalized therapy” section. 2.2. Mechanisms of Cell Death in BNCT In BNCT, cells are damaged mainly by alpha particles or the 7 Li ion, as they can cause various DNA lesions along their path, i.e., DNA damage in clusters or multiple local damage sites, resulting in genome instability. In addition to DNA, macromolecules can also be damaged, resulting in modulation of their functions [ 38 ]. When these particles pass through a cell, their path is short ( α < 10 µ m and 7 Li < 5 µ m) [ 39 ], so their kinetic energy is released within the target cell, whose diameter is usually ~10 µ m. Therefore, it does not affect the surrounding healthy cells. Intracellular boron localization is critical because normal healthy tissue can be spared from nuclear reactions if it does not uptake 10 B. Unfortunately, with the currently available boron carrier compounds, some 10 B also accumulate in healthy cells. The development of boron carriers is still highly active today [ 40 ]. A schematic representation of the principles of action at cellular level is presented in Figure 3. Cancers 2022, 14, x 6 of 26 boron-dispersed nanoparticles [31–34] (nanomaterial-based delivery systems), and others currently under evaluation. Targeted boron delivery agents combine boron-containing agents with tumor-targeting molecules (e.g., nucleosides, porphyrins, peptides, proteins, or antibodies). Boron-delivery nanomaterials can transport various boron-containing compounds into tumor cells by taking advantage of the enhanced permeability and retention effect that affects nanomaterials and the active targeting effects mediated by tumortargeted ligands grafted on the surface of the materials. The following boron-delivery nanomaterials can be distinguished: dendrimers, liposomes, polymeric nanoparticles, boron nitride, carbon nanotubes, mesoporous silica nanoparticles, ferromagnetic and paramagnetic nanoparticles—some of them used for MRI imaging—, gold nanoparticles, and BPO4 nanoparticles. There are many promising routes in drug delivery systems, and there is still a pressing need to develop new boron delivery agents, but without adequate research and clinical trials, it is difficult to determine which is the most feasible [35,36]. There are some obstacles to the exploitation of the full potential of the cancer-specific selectivity of BNCT, including the suboptimal boron delivery strategies currently used. However, various supportive targeted measures are under development, for example, delivery agents targeting the glucose transporter GLUT1 [37]. In any case, BNCT workflow would allow for a full personalization of the treatment, from the drug delivery treatment to imaging, and considering high throughput techniques, as it is discussed later in the “BNCT and personalized therapy” section. 2.2. Mechanisms of Cell Death in BNCT In BNCT, cells are damaged mainly by alpha particles or the 7Li ion, as they can cause various DNA lesions along their path, i.e., DNA damage in clusters or multiple local damage sites, resulting in genome instability. In addition to DNA, macromolecules can also be damaged, resulting in modulation of their functions [38]. When these particles pass through a cell, their path is short (α < 10 µm and 7Li < 5 µm) [39], so their kinetic energy is released within the target cell, whose diameter is usually ~10 µm. Therefore, it does not affect the surrounding healthy cells. Intracellular boron localization is critical because normal healthy tissue can be spared from nuclear reactions if it does not uptake 10B. Unfortunately, with the currently available boron carrier compounds, some 10B also accumulate in healthy cells. The development of boron carriers is still highly active today [40]. A schematic representation of the principles of action at cellular level is presented in Figure 3. Figure 3. Schematic representation of BNCT principles of action at a cellular level. Boron accumulation is favored in tumoral cells (thick upper arrow) in comparison with normal cells (thin upper arrow). Figure 3. Schematic representation of BNCT principles of action at a cellular level. Boron accumulation is favored in tumoral cells (thick upper arrow) in comparison with normal cells (thin upper arrow). Radio-induced damage can be produced through two types of action: direct and indirect. In the first case, radiation directly affects DNA, causing the ionization of atoms within the DNA molecule in small fractions of seconds. However, radiation-caused ionization must take place within a few nanometers of the DNA molecule to directly damaging it. As with indirect damage, radiation interacts with other target molecules or atoms that it encounters, usually water [ 41 ]. As a result, highly reactive species, such as HO and H, are Cancers 2022,14, 2865 7 of 26 produced and they can diffuse some distance in the cell before reaching the place where damage will be produced. Different types of DNA injury include base damage, DNA protein or DNA–DNA crosslinks, double-strand breaks (DSB), single-strand breaks (SSB), sugar-phosphate backbone interruption, etc. Their distribution and repair pathways depend strongly on the type of radiation used during BNCT and its LET characteristics [11,42]. DNA damage generally increases together with radiation LET [ 43 ], and the higher the LET, the higher the relative biological effectiveness (RBE). Takatsuji et al., discussed the relationship between LET and RBE considering chromosomal aberrations and cell death, and found that at low doses, RBE increases with increasing LET, then the RBE value peaks at a LET of about 100 keV ·µ m −1 , and, finally, the RBE decreases as LET increases further [ 44 ]. The radiation field generated during BNCT consists of components with different LET characteristics that act independently. Low-LET radiation ionizes sparsely, whereas high-LET radiation causes denser ionization along the track and can lead to more complex DNA damage. DNA damage after high-LET radiation remains unrepaired for a long time, leading to genome instability or cell death [45]. The density of radiation affects the presence and quality of radiation induced DSB. Like many cancer treatments, radiation therapy achieves its therapeutic effect by causing a reaction of several types of cell death: apoptosis, mitotic cell death or mitotic catastrophe, necrosis, autophagy, and others. Apoptosis or mitotic cell death are the most common types, along with necrosis. Radiation-induced apoptosis is a progressive and degrading process. Extrinsic (death receptor) and intrinsic (mitochondrial) apoptosis can be distinguished. Intrinsic apoptosis is a type of regulated cell death (RCD) that is initiated by perturbations of the intracellular microenvironment and marked by permeabilization of the mitochondrial outer membrane. The extrinsic apoptotic pathway is triggered by binding of death ligands to transmembrane receptor proteins (e.g., TNFα to TNFR1). Mitotic cell death is a specific variant of RCD driven by mitotic catastrophe, an oncosuppressive mechanism to control mitosis-incompetent cells [ 46 ]. Wang et al. [ 47 ] confirmed that in glioma cells, BNCT-induced apoptosis was mediated by the Bcl-2/Bax pathway. Another important goal of radiation therapy is to deprive cancer cells of their potential to divide and multiply indefinitely [ 7 ]. The primary and presumed cell target of the ionizing radiation is DNA itself. However, damage or mutations in different cellular macromolecules cannot be fully eliminated, and as a result, their functions could be modulated, and other subsequent biological changes can be observed after cancer treatment [38]. The DNA-DSB repair process is complex and depends on many factors, including the cell cycle phase and checkpoints, DSB-inducing agents, non-coding RNAs, and various gene mutations characterized by different cancer cell lines. Several attempts have been made over the past few years to investigate the specific response to cellular DNA damage induced by a mixed neutron-gamma field [ 45 , 48 , 49 ]. Despite that, this phenomenon is not fully understood and determined. Rodriguez et al. [ 50 ] have attempted in vitro studies of DNA damage and repair mechanisms induced by BNCT. The human thyroid follicular cancer cell line was used for the research. DNA damage assessment was performed by detecting H2AX histone phosphorylation foci ( γ H2AX foci). Repair of DBS through two mediating pathways has been identified in mammalian cells, homologous recombination repair (HRR) and nonhomologous end joining (NHEJ). After the analysis of the follicular carcinoma cell analysis, the repair pathways were observed with an increase in Rad51 and Rad54 mRNA expression 4 and 6 h after irradiation, showing the expression of enzymes that belong mainly to the HRR pathway, specifying a different pattern of DNA damage and showing activation of both repair pathways. However, what exactly determines the activation of HRR or NHEJ is not yet completely clear. To increase anticancer biological activity during BNCT therapy, Ikuhiko Nakase et al., performed an in vitro BNCT assay [ 26 ]. The study also evaluated cell death pathways to understand the cell killing activity that occurs after thermal neutron irradiation. They syn- Cancers 2022,14, 2865 8 of 26 thesized and demonstrated organelle-targeted cell-penetrating peptide (CPP)-conjugated boron compounds. CPPs help to control intracellular localization, cell membrane penetration, and further enhance cellular uptake of the boron compound. This controlled delivery affects the types of cell death and the efficacy of cancer cell killing activity. Treatment with DB-RLA ((BODIPY)-labeled dodecaborates conjugated to an RLA peptide) showed a higher reduction in the ATP content than other peptides tested. ATP depletion enhances necrosis, which consequently might induce necrosis in BNCT. This could be one of the significant factors of the cell-killing activity and detailed mechanisms should be further studied. 2.3. Physical Basis and Dosimetry of BNCT Ionizing radiation has many forms, from alpha, beta, proton, or neutron particles, to X or gamma rays, and others. The main components that contribute to the total absorbed dose rate in BNCT are the elastic interaction of incident neutrons with hydrogen, the gamma ray dose emitted by the source, and the thermal neutrons captured by hydrogen, nitrogen, and boron [ 51 ]. Each of the components has various biological weighting factors. The total biologically absorbed dose (Gy-Eq) is the sum of the physical dose components (D) multiplied by the compound biological effectiveness (CBE) or relative biological effectiveness (RBE) of each dose component. RBE is the ratio of the dose absorbed from the reference radiation to the value of the radiation dose tested, producing the same biological effect, whereas CBE represents values of the biological efficacy for each dose component, depending on the boron carrier used. CBE differs from RBE with high LET general radiation in that the value varies depending on target cells, tissue, and the type of boron compounds used [ 52 ]. However, because of the occurrence of events at the cellular and subcellular levels, the different energies and types of radiation involved, the dosimetry, and accurate estimation of RBE, CBE, and, therefore, the biological effectiveness of BNCT is challenging. Streitmatter et al. [ 53 ] presented a multiscale system of dosimetric and radiobiological models that better assess biological effectiveness. It can predict not only CBE and RBE but also other critical biological metrics for neutron sources, such as boron micro-distribution and tissue types. The model was tested against results from published experiments in vitro and in vivo, with and without boron, and showed good agreement between both. As the biodistribution of boron varies from patient to patient, determining the boron concentration, as will be discussed in Section 3, is one of the crucial factors often marked as a drawback in BNCT because it causes ambiguity in the calculated dose distributions. For effective treatment with BNCT, the ratio of 10 B concentration in the tumor and its concentration in normal tissues (T/N ratio) should be 3:1 or more, and the concentration of 10 B in the target should be at least ∼ 15–30 µ g g −1 or ∼ 10 9 atoms per cell to perform lethal tumor cell damage [ 54 ]. In summary, to avoid unfavorable effects, the concentration of 10 B in tumor cells and normal tissues must be known to support the calculation of the total dose distribution and to allow a good prognosis by allowing appropriate patient selection and optimization based on the spatial distribution of the boron-containing compound known before treatment. Human tissue also contains certain isotopes that react with neutrons. Due to the values of nuclear cross sections, the most meaningful interactions of neutrons with human tissue involve 1 H, 12 C, 14 N, and 16 O isotopes, which account for 99.2% of all atoms in the human body [ 55 ]. Consequently, all the described components should be considered during the evaluation and calculation of the doses received because they could also be responsible for the adverse effects of BNCT. The most advanced methods of calculating fluxes and doses in complex geometries with a heterogeneous physical density, such as those of the patient are based on Monte Carlo (MC) techniques. This mathematical method uses the probability distribution that describes the transport of particles to determine the outcome of each step of their history along the way through the mediums by randomly sampling. In this way, you can know the position, direction, and energy, among other variables, of each particle in the radiation fluxes. Today, it is accepted that the probability description of particle transport is accurate enough with the energy values involved, so the MC method is Cancers 2022,14, 2865 9 of 26 the gold standard tool for dose calculation or design of new prototypes for the generation of neutron sources. Before a clinical application, BNCT, like any other radiotherapeutic technique, requires a preceding dose calculation to determine the proper operation of the device and the irradiation condition by considering all design features and beam shaping components with respect to material, motion, and geometry for dose delivery. These processes are carried out by the treatment planning systems (TPS) [ 56 ]. Despite the high time consuming inherent to the Monte Carlo method, this shows a high accuracy compared to the other analytic algorithms. Thus, most BNCT TPS use a MC method to estimate the absorbed dose in some part of the whole process. Many different tools and dose calculation algorithms based on the MC method have been developed and are being developed continuously that can assist clinicians in personalized treatment planning and decision making. Nowadays, the dose distribution and mean doses absorbed in regions of interest (ROI), and the dose volume histograms and isodose curves superimposed on personalized anatomical models of the patients can be extracted and displayed graphically after appropriate data calculation. The shorter the recording step to track the transport of particles along the material media described in the image, the more accurate MC dose calculation is, but a smaller registration volume for the calculation of the average dose value relates to higher statistical uncertainty due to a lower number of events in the volume. Therefore, the computation time is an important handicap for the clinical application of MC. Some attempts have been made to improve the MC algorithms, e.g., by increasing the dose calculation speed of BNCT TPS, such as in GPU-accelerated Monte Carlo [ 57 ]. Although some companies are working on a commercial MC dose engine focused specifically on the clinical application of BNCT, the MC method is often used as an alternative tool to verify specific commercial TPS [ 58 ]. The general-purpose MC radiation transport code MCNP [ 59 ] has been used recurrently as a dose calculation engine in BNCT. Historically, the ‘Simulation Environment for Radiotherapy Application’ (SERA) [60] has been used in BNCT facilities using a special MC calculation engine. On the other hand, more recently, Hu et al., evaluated the Particle and Heavy Ion Transport code System (PHITS) [ 61 ], a multimodal Monte Carlo code developed by the Japan Atomic Energy Agency in Japan, for micro dosimetry in BNCT [ 62 ]. It can help to evaluate doses in radiobiological experiments. In addition, it can consider intracellular and intercellular heterogeneity in the 10 B distribution. Therefore, it was proposed as a model that can estimate the biological effectiveness of newly developed 10 B compounds for BNCT, which would be advantageous in future drug discovery research. The study resulted in the general conclusion that PHITS can be applied to evaluate the dose rates of absorbed gamma rays and the thermal neutron fluxes within a tumor-imitating medium [62,63]. In the clinical dose calculation, several components of the absorbed dose must be considered. Four main components of the absorbed dose in BNCT can be distinguished: 1. Fast neutron dose: according to 1 H(n, n)preactions, fast and epithermal neutrons cause elastic neutron collisions with hydrogen in tissue (giving recoiling protons and gammas). Other energy depositions from fast neutron reactions like 12 C(n, α ) are also included. 2. Incident and secondary gamma ray’s dose: Primary gamma dose from the beam port and secondary gamma dose by 1 H (n, γ ) 2 H. This component can be used for real time dosimetry with SPECT imaging, as it is further described in Section 3. 3. Nitrogen dose: according to the 14 N (n,p) 14 C reaction, the 14 N element in the tissue captures a thermal neutron and, as a result, a ~600 keV proton is emitted. The dose is obtained from locally delivered energy from the recoiling 14 C nucleus and the energetic proton. 4. Boron dose: energy deposited by the 10 B (n, α ) 7 Li reaction. 10 B captures a thermal neutron, and as a result, an alpha particle and a recoiling 7 Li ion are emitted. The dose derived from the reaction products is ~2.31 MeV. The cross sections for some of these reactions are shown in Figure 4. Cancers 2022,14, 2865 16 of 26 reactor sources. With the improvement of neutron beam-generating instruments, BNCT would be able to improve the robustness of the clinical trial results by increasing the number of patients included in these studies. BNCT has been the subject of several clinical trials over recent decades. Cancers 2022, 14, x 16 of 26 [115]. Within Europe, Neutron Therapeutics designed one of the first commercial accelerator-based Boron Neutron Capture Therapy Platforms and installed it in the Helsinki University Hospital. This facility is the base for the process of obtaining the European CE safety certification for medical devices, necessary for their commercialization. This process is currently ongoing there. TAE Life Sciences (TLS) is a company for biologically targeted radiation therapy developing a breakthrough accelerator-based BNCT system, which has formalized an agreement with the National Center of Oncological Hadron-therapy (CNAO) in Italy to provide its Alphabeam™ Neutron System, which integrates RayStation of RaySearch Laboratory, the more extended software for treatment planning in RT. The latter is a clear signal of the new interest in BNCT, despite being a fairly old technique, due to the recent generation of these new compact devices that could treat patients in conditions similar to those of other particle accelerators in hospital settings. Figure 5. On the left side, the Sumitomo BNCT System NeuCure was recently approved by the Japanese government and is running in two hospitals from early 2020. Image adapted from [116]. On the right side, a compact accelerator of Proteus®ONE protons, installed at Willis-Knighton Center in Los Angeles, USA, clinically used from 2014. Image adapted from [117]. 4.2. BNCT Clinical Trials Until recently, the value of BNCT was largely restricted and the number of patients treated with BNCT was limited because treatment could only be performed in nuclear research reactors—the only neutron source at the time—as previously discussed in Section 4.1. Furthermore, most of the clinical trials conducted were carried out in facilities at nuclear reactor sources. With the improvement of neutron beam-generating instruments, BNCT would be able to improve the robustness of the clinical trial results by increasing the number of patients included in these studies. BNCT has been the subject of several clinical trials over recent decades. The first clinical trials date back to the 1950s [23,24] using first-generation compounds with poor biodistribution, which was the main cause that led to their failure, as previously discussed [118–120]. Some of the first reported clinical trials using second-generation boron carriers date back to the late 1980s/early 1990s [121]. Several locations have been key in the development of BNCT and its clinical trials, being one of them the Finnish Research Reactor (FiR 1 reactor), where some clinical trials started at 1999. In 2003, Joensuu et al., summarized the ongoing clinical trials at these facilities: one trial for glioblastoma patients who had not undergone surgery or radiotherapy before BNCT, and another for recurring or progressing glioblastoma patients who had previously undergone surgery or conventional cranial radiotherapy. Both phase I/II Figure 5. On the left side, the Sumitomo BNCT System NeuCure was recently approved by the Japanese government and is running in two hospitals from early 2020. Image adapted from [ 116 ]. On the right side, a compact accelerator of Proteus ® ONE protons, installed at Willis-Knighton Center in Los Angeles, USA, clinically used from 2014. Image adapted from [117]. The first clinical trials date back to the 1950s [ 23 , 24 ] using first-generation compounds with poor biodistribution, which was the main cause that led to their failure, as previously discussed [ 118 – 120 ]. Some of the first reported clinical trials using second-generation boron carriers date back to the late 1980s/early 1990s [121]. Several locations have been key in the development of BNCT and its clinical trials, being one of them the Finnish Research Reactor (FiR 1 reactor), where some clinical trials started at 1999. In 2003, Joensuu et al., summarized the ongoing clinical trials at these facilities: one trial for glioblastoma patients who had not undergone surgery or radiotherapy before BNCT, and another for recurring or progressing glioblastoma patients who had previously undergone surgery or conventional cranial radiotherapy. Both phase I/II clinical trials concluded that BNCT was well tolerated and established the foundations for fructose-BPA-based BNCT applications in the clinic [ 122 ]. Later, in 2007, Kankaanranta et al., published the results of another Finnish trial that evaluated BNCT as a treatment for locally recurrent head and neck cancer in another phase I/II clinical trial with two irradiation sessions. The results showed that BNCT was effective and safe to use in patients with previously irradiated head and neck cancer who had recurrences [ 123 ]. Almost five years later, in early 2012, a final analysis revealed that most of the patients were BNCT respondents, some of them were progression-free for sustained periods, and only one patient of the cohort progressed in their disease stage. They also reported acceptable toxicity levels [ 124 ]. Additionally, in 2011, Kankaanranta et al., presented the results of a clinical trial evaluating BNCT in malignant gliomas resistant to surgery and conventional radiotherapy. The main objective of this trial was to determine the biosafety of different doses of BPA ranging from 290 mg/kg to 450 mg/kg, which was determined as the maximum tolerated dose in which some adverse effects appeared. Furthermore, they estimated that irradiation doses were higher in patients administered with more than 290 mg/kg of fructose-BPA compared to those administered only 290 mg/kg. Therefore, they concluded that BPA-based BNCT with doses of up to 400 mg/kg was a feasible treatment for malignant recurrent gliomas [ 125 ]. Cancers 2022,14, 2865 17 of 26 The data from the patients in this study was used afterwards to formulate and train a pharmacokinetic model of boron biodistribution in glioma patients [126]. In the UK, in 2009, Cruickshank et al., reported preliminary data from a clinical trial started in 2007 to evaluate BPA pharmacokinetics for patients with glioma. More specifically, they worked on the location of the intravenous infusion and the use of mannitol as a controlled blood–brain barrier disrupter, and its effect on boron biodistribution. However, this trial was terminated and no further information was found. The only information shared was from a single patient infused without mannitol, whose biodistribution data were consistent with data from Finnish trials [127]. In late 2016, Yong et al., reported the outcome of their first case in the clinical trial as the preliminary results of the In-Hospital Neutron Irradiator for treating malignant melanoma with BNCT in China. In this phase I/II clinical trial, BPA-fructose was used at a dose of 350 mg/kg infused intravenously for 90 min and the concentration of circulating boron was measured over time by performing several blood extractions of the patient. Irradiation was carried out successfully and the patient showed a complete response to BNCT without late radiation injury and only some minor grade 2 acute radiation damage that resolved after proper treatment [128]. In Taiwan, a phase I to II BNCT clinical trial was conducted between 2010 and 2013 for patients with recurrent head and neck cancer. They were treated with a two-session BNCT treatment using fructose BPA, and reported a wide range of responses, including a complete response in half of the patients, and partial responses with various reactions, some of which were disease-free for more than 50 months after BNCT [ 129 ]. A second clinical trial, also phase I/II, was started in 2014 combining BNCT with IGRT and IMRT to try to reduce re-recurrences [ 130 ]. As of 2019, Lee et al., provided evidence suggesting that the single-session dose distribution in terms of homogeneity and conformity might improve when combined with multifraction IMRT [ 131 ]. Additionally, in Taiwan, BNCT was evaluated as a salvage therapy for malignant brain tumors, with preliminary results published in 2020 showing significant effects on maintaining a good quality of life and possibly prolonging a patient’s survival [ 132 ]. Their latest results were published in 2021, showing no severe adverse effects, a possible increase in survival, and also establishing some key parameters and recommendations that patients should meet to undergo BNCT or at least the conditions under which BNCT showed better results [133]. At the time of writing this review, there is one open and recruiting BNCT-related clinical trial listed in the ClinicalTrials database with reference NCT04293289. This study based in Japan is a phase I clinical trial of BNCT for malignant melanoma and angiosarcoma using an experimental CICS-1 BNCT accelerator and SPM-011 (borofalan) intravenously at 200 mg/kg/h for 2 h before neutron irradiation and a continuous infusion at 100 mg/kg/hour during irradiation. This trial started in late 2019 and is expected to be finished by the end of 2022. Although these trials have been steadily increasing, the lack of phase III trials comparing the effectiveness of BNCT against conventional radiation treatment is a flaw to overcome in the coming years, as well as fully exploring the features provided by BNCT towards a more patient-specific approach. 4.3. BNCT and Personalized Radiotherapy RT in general is searching for new methodologies for rational patient treatment based on molecular knowledge and involving imaging in radiobiology to assess the therapeutic outcome of targeted drugs in combination with radiation, in addition to the toxicities that limit the escalation of prescribed dose of the treatments. As discussed previously, in recent decades, radiotherapy has experienced significant advances, mainly due to technological progress. These advances led to prominent levels of precision in delivering to the tumor volume a prescription dose based on population evidence. However, new research lines are trying to find treatment plan and a prescribed dose specific for each patient once delivery precision has been assured. Cancers 2022,14, 2865 18 of 26 In this sense, it is necessary to characterize the biological reality of the tumor, and, hence, the latest developments in radiotherapy are related to its personalization at a molecular level. At this point, both omic techniques and functional imaging are the key approaches used to make even more personalized radiotherapy, integrating patientspecific molecular data with population-based data, such as clinical endpoints and toxicity constraints [ 134 ]. Examples of such personalization are the use of radiotracers that label hypoxia levels at a treatment planning level, contouring, and prescribing doses based on zones with different proliferation levels, and even using RT as an immunotherapy adjuvant for oligometastatic disease [ 135 – 139 ]. The personalization level has been brought even to prescribing doses to different tumoral regions identified in functional imaging that obey different radiation resistance levels and/or to the true pathoanatomical borders of the tumor. This specific dose prescription based on the heterogeneity provided by the molecular image has been proposed even up to individual voxels corresponding to the tumor volume after the appropriate segmentation process in the image [ 140 ]. This approach has been named dose painting [ 141 ] and is being slowly implemented in treatment planning because harmonization protocols must still be established previously between a larger number of image devices worldwide, specifically PET/CT [ 142 ], enough to achieve common quantitative metrics from a larger cohort, thus being able to confirm the robust correlation between the gray level in the image and the biological information sensible to the prescription dose decision in each case. As Ree et al. argued, considering these biological and molecular characteristics can contribute to selective sensitization of the tumor for optimization of tumor control, reducing normal tissue injury from radiation damage, and even modulating the immune system for better management of locally advanced, and possibly also metastatic disease [134]. The same principles apply to BNCT, where direct personalization could be achieved through selective molecular imaging based on boron biodistribution for subsequent highly precise targeted radiotherapy planning, as discussed in Section 2. Nevertheless, approaches based on high throughput omics will also be desirable. As Sauerwein et al., discussed in early 2021, BNCT personalization is based on the 10 B targeting uptake mechanism, tunable using different vectors based on previously determined molecular characteristics of the tumor. This uptake would affect the radiation dose delivered at the cellular and even subcellular levels, but more importantly, this boron uptake and biodistribution will also depend on the characteristics of each patient from a pharmacogenetic point of view [ 36 ]. Therefore, several models have been developed to evaluate and predict the pharmacokinetics and irradiation effect of BNCT using different molecular ligands in boron transporters, such as the in vitro model proposed by Ishiyama et al. [ 136 ] using three-dimensional tumoral human cell cultures. Regardless, focusing on proper molecular profiling using high throughput approaches is a fundamental step that needs to be tackled in the near future for more complete personalization of BNCT. These studies should not only be limited to genomics or transcriptomics, but also include proteomics or metabolomics to have a clearer picture of the problem, identify and classify the different stages of the disease more effectively, and avoid the false static and limited view that can provide genetic approaches. Experimental studies also evaluated the relationship between tumor temperature and the response to BNCT application. The main objective was to measure temperature before irradiation and to check if it could be used as a predictive indicator of boron tumor uptake, so BNCT could be personalized and optimized for each individual patient by adjusting neutron fluence [143]. Few omic approaches for BNCT have been reported. Some of the first procedures related to BNCT proteomics were those discussed by Mauri and Basilico, in 2012, where the focus of the study was to determine the interaction of boron transporters with endogenous proteins, using techniques, such as bidimensional gel electrophoresis and multidimensional protein identification [144]. Cancers 2022,14, 2865 19 of 26 Later, in 2015, Sato et al., studied the cellular response to BNCT in SAS human squamous cell carcinoma, with and without previous incubation with BPA. Their findings showed that the proteome presented alterations related to processes, such as DNA repair and RNA processing, as well as changes in proteins located in the cellular compartment of the endoplasmic reticulum. Thus, such proteins could be involved in the early response to BNCT. More precisely, they found that lymphoid-restricted membrane proteins were induced after BNCT and may be related to BNCT-induced cell death [83]. In 2019, Ferrari et al. [ 84 ] reported a proof-of-concept study in which urinary samples were used to characterize the proteome of patients with squamous cell head and neck cancer and thyroid cancer in a BNCT clinical trial. Samples were taken before and after the administration of the boron agents. Several candidate biomarkers were found and changes in the proteome were detected after boron infusion, more precisely the reduction in the expression levels of three inflammation-related molecules. In any case, as Keener pointed out in late 2020, whether through genetics or advanced imaging, the field of radiation oncology is slowly but steadily adopting the principles of personalized medicine [145] and BNCT shows a potential higher efficiency for clinical implementation since it provides cellular targeting of energy transfer radiation and, therefore, can provide more useful previous genetic and biological information. 5. Conclusions Boron Neutron Capture Therapy incorporates the specific principles of some chemotherapies and targeted therapies into the precise location principles of conventional radiotherapy. In this way, BNCT could share the typical methodologies used in personalized medicine. Since some types of cancer, such as glioblastoma, remain exceptionally resistant to all current forms of therapy, such as chemotherapy, surgery, radiotherapy, and immunotherapy, BNCT is a promising option for these types of tumors. However, some critical issues must be resolved if BNCT is to become a better and more valuable cancer treatment. Since controlled intracellular targeting is of great importance in inducing the cell-killing activity of BNCT due to specific cell death pathways, such targeting should be further assessed, together with the conduct of adequate research and clinical trials to determine the most profitable and promising routes in drug delivery systems. Activation of the DNA response, such as damage and repair mechanisms of complex double-strand DNA break activated by a mixed neutron-gamma beam, has been poorly studied and, therefore, it is not fully determined. A deeper understanding of how cells preferentially select specific DNA damage responses generated by high-LET and mixed radiation, and detailed mechanisms of enhanced necrosis due to ADP depletion, can lead to improved therapeutic efficiency in BNCT. Individual tumor cell quantification of bound and free pools (net cellular content) of 10 B needs to be further addressed, as it remains challenging (owing to insufficient spatial resolution) with clinically applicable techniques. Studies designed to test and improve boron detection methods could reduce detection limits and identify accumulation regions in tumor cells and normal tissues with greater precision. Further research on mechanisms for detecting the distribution of prompt gamma rays that arise during BNCT could also be profitable. However, the ideal dose paradigm for BNCT, the real-time measurement of the distributions of reactions, such as 10 B(n, α ) 7 Li and 14 N(n,p) 14 C, and the quantitative mapping of the boron concentration in the body have not yet been determined. Finally, the applicability of the BNCT boron-containing drug used in the individual patient must be evaluated for effective treatment, not only by PET and other individual imaging methods, but also by proteomics and gene expression methods, as is beginning to be performed for BPA (Borofalan). Cancers 2022,14, 2865 20 of 26 Author Contributions: Conceptualization, A.L., M.B.; Data curation, A.L., D.S., J.A.L.-V., M.B.; Funding acquisition, A.L.; Investigation A.L., D.S., J.A.L.-V., M.B.; Methodology, A.L., D.S., J.A.L.-V., M.B.; Project administration, A.L.; Supervision, A.L., M.B.; Validation, A.L., D.S., J.A.L.-V., M.B.; Visualization A.L., D.S., J.A.L.-V., M.B.; Writing—original draft, A.L., D.S., J.A.L.-V., M.B.; Writing— review & editing. A.L. and M.B. were involved in study design, manuscript style, and critically reviewed the text by contributing expertise and guidance. A.L. leads the scientific projects by which this work is supported. All authors have read and agreed to the published version of the manuscript. Funding: Grant RTI2018-098656-B-100 funded by MCIN/AEI/ 10.13039/501100011033 and by “ERDF A way of making Europe”, by the “European Union”. The APC was also funded by this research project. This research was also possible thanks to Project P20_01053 funded by the European Union and the Junta de Andalucía through the European Regional Development Fund (FEDER); D.S. received funding from ERASMUS PLUS-European Commission program for 2014–2020, project number 2019-1-PL01-KA103-062953; J.A.L.V. received funding from the FPU call from the Spanish Ministry of Science, Innovation, and Universities, reference number FPU19/02950. Conflicts of Interest: The authors declare no conflict of interest. References 1. Sung, H.; Ferlay, J.; Siegel, R.L.; Laversanne, M.; Soerjomataram, I.; Jemal, A.; Bray, F. Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries. CA A Cancer J. Clin. 2021 ,71, 209–249. [CrossRef] [PubMed] 2. Hanahan, D.; Weinberg, R.A. The Hallmarks of Cancer. Cell 2000,100, 57–70. [CrossRef] 3. Hanahan, D.; Weinberg, R.A. Hallmarks of Cancer: The Next Generation. Cell 2011,144, 646–674. [CrossRef] [PubMed] 4. Atun, R.; Jaffray, D.A.; Barton, M.B.; Bray, F.; Baumann, M.; Vikram, B.; Hanna, T.P.; Knaul, F.M.; Lievens, Y.; Lui, T.Y.M.; et al. Expanding Global Access to Radiotherapy. Lancet Oncol. 2015,16, 1153–1186. [CrossRef] 5. Baskar, R.; Lee, K.A.; Yeo, R.; Yeoh, K.-W. Cancer and Radiation Therapy: Current Advances and Future Directions. Int. J. Med. Sci. 2012,9, 193. [CrossRef] 6. Fiorino, C.; Guckemberger, M.; Schwarz, M.; van der Heide, U.A.; Heijmen, B. Technology-driven Research for Radiotherapy Innovation. Mol. Oncol. 2020,14, 1500. [CrossRef] 7. Chen, Y.; Yang, J.; Fu, S.; Wu, J. Gold Nanoparticles as Radiosensitizers in Cancer Radiotherapy. Int. J. Nanomed. 2020 ,15, 9407. [CrossRef] 8. Farhood, B.; Samadian, H.; Ghorbani, M.; Zakariaee, S.S.; Knaup, C. Physical, Dosimetric and Clinical Aspects and Delivery Systems in Neutron Capture Therapy. Rep. Pract. Oncol. Radiother. 2018,23, 462–473. [CrossRef] 9. Nedunchezhian, K.; Aswath, N.; Thiruppathy, M.; Thirugnanamurthy, S. Boron Neutron Capture Therapy—A Literature Review. J. Clin. Diagn. Res. 2016,10, ZE01–ZE04. [CrossRef] 10. Macías, M.; Fernández, B.; Praena, J. New Data for the Definition of Neutron Beams for Boron Neutron Capture Therapy. Radiat. Phys. Chem. 2020,185, 109474. [CrossRef] 11. Dymova, M.A.; Taskaev, S.Y.; Richter, V.A.; Kuligina, E.V. Boron Neutron Capture Therapy: Current Status and Future Perspectives. Cancer Commun. 2020,40, 406–421. [CrossRef] [PubMed] 12. Espain, M.S.; Dattoli Viegas, A.M.; Trivillin, V.A.; Saint Martin, G.; Thorp, S.I.; Curotto, P.; Pozzi, E.C.C.; González, S.J.; Portu, A.M. Neutron Autoradiography to Study the Microdistribution of Boron in the Lung. Appl. Radiat. Isot. 2020,165, 109331. [CrossRef] [PubMed] 13. Wittig, A.; Michel, J.; Moss, R.L.; Stecher-Rasmussen, F.; Arlinghaus, H.F.; Bendel, P.; Mauri, P.L.; Altieri, S.; Hilger, R.; Salvadori, P.A.; et al. Boron Analysis and Boron Imaging in Biological Materials for Boron Neutron Capture Therapy (BNCT). Crit. Rev. Oncol./Hematol. 2008,68, 66–90. [CrossRef] [PubMed] 14. Qi, P.; Chen, Q.; Tu, D.; Yao, S.; Zhang, Y.; Wang, J.; Xie, C.; Pan, C.; Peng, H. The Potential Role of Borophene as a Radiosensitizer in Boron Neutron Capture Therapy (BNCT) and Particle Therapy (PT). Biomater. Sci. 2020,8, 2778–2785. [CrossRef] [PubMed] 15. Wang, D.; Meng, Y.; Wang, X.; Xia, G.; Zhang, Q. The Endocytic Mechanism and Cytotoxicity of Boron-Containing Vesicles. Chem. Pharm. Bull. 2020,68, c10-00971. [CrossRef] 16. Miyatake, S.-I.; Kawabata, S.; Hiramatsu, R.; Kuroiwa, T.; Suzuki, M.; Ono, K. Boron Neutron Capture Therapy of Malignant Gliomas. Prog. Neurol. Surg. 2018,32, 48–56. [CrossRef] 17. Yokoyama, K.; Miyatake, S.-I.; Kajimoto, Y.; Kawabata, S.; Doi, A.; Yoshida, T.; Okabe, M.; Kirihata, M.; Ono, K.; Kuroiwa, T. Analysis of Boron Distribution In Vivo for Boron Neutron Capture Therapy Using Two Different Boron Compounds by Secondary Ion Mass Spectrometry. Radiat. Res. 2007,167, 102–109. [CrossRef] 18. Puris, E.; Gynther, M.; Auriola, S.; Huttunen, K.M. L-Type Amino Acid Transporter 1 as a Target for Drug Delivery. Pharm. Res. 2020,37, 88. [CrossRef] 19. CSID:132497 L-BPA|C9H12BNO4|Chemspider. Available online: http://www.chemspider.com/Chemical-Structure.132497.html (accessed on 1 June 2022). Cancers 2022,14, 2865 21 of 26 20. Hansch, C.; Rockwell, S.D.; Jow, P.Y.C.; Leo, A.; Steller, E.E. Substituent Constants for Correlation Analysis. J. Med. Chem. 1977 ,20, 304–306. [CrossRef] 21. Suzuki, M. Boron Neutron Capture Therapy (BNCT): A Unique Role in Radiotherapy with a View to Entering the AcceleratorBased BNCT Era. Int. J. Clin. Oncol. 2020,25, 43–50. [CrossRef] 22. Borofalan (10B)—Stella Pharma—AdisInsight. Available online: https://adisinsight.springer.com/drugs/800045201 (accessed on 16 September 2021). 23. Farr, L.E.; Sweet, W.H.; Robertslon, J.S.; Foster, C.G.; Locksley, H.B.; Sutherland, D.L.; Mendelsohn, M.L.; Stickley, E.E. Neutron Capture Therapy with Boron in the Treatment of Glioblastoma Multiforme. Am. J. Roentgenol. Radium. Ther. Nucl. Med. 1954 ,71, 279–293. [PubMed] 24. Goodwin, J.T.; Farr, L.E.; Sweet, W.H.; Robertson, J.S. Pathological Study of Eight Patients with Glioblastoma Multiforme Treated by Neutron-Capture Therapy Using Boron 10. Cancer 1955,8, 601–615. [CrossRef] 25. Hoppenz, P.; Els-Heindl, S.; Kellert, M.; Kuhnert, R.; Saretz, S.; Lerchen, H.-G.; Köbberling, J.; Riedl, B.; Hey-Hawkins, E.; Beck-Sickinger, A.G. A Selective Carborane-Functionalized Gastrin-Releasing Peptide Receptor Agonist as Boron Delivery Agent for Boron Neutron Capture Therapy. J. Org. Chem. 2020,85, 1446–1457. [CrossRef] [PubMed] 26. Nakase, I.; Katayama, M.; Hattori, Y.; Ishimura, M.; Inaura, S.; Fujiwara, D.; Takatani-Nakase, T.; Fujii, I.; Futaki, S.; Kirihata, M. Intracellular Target Delivery of Cell-Penetrating Peptide-Conjugated Dodecaborate for Boron Neutron Capture Therapy (BNCT). Chem. Commun. 2019,55, 13955–13958. [CrossRef] 27. Nakase, I.; Aoki, A.; Sakai, Y.; Hirase, S.; Ishimura, M.; Takatani-Nakase, T.; Hattori, Y.; Kirihata, M. Antibody-Based Receptor Targeting Using an Fc-Binding Peptide-Dodecaborate Conjugate and Macropinocytosis Induction for Boron Neutron Capture Therapy. ACS Omega 2020,5, 22731–22738. [CrossRef] 28. Imperio, D.; Muz, B.; Azab, A.K.; Fallarini, S.; Lombardi, G.; Panza, L. A Short and Convenient Synthesis of Closo-Dodecaborate Sugar Conjugates. Eur. J. Org. Chem. 2019,2019, 7228–7232. [CrossRef] 29. Cappelli, A.; Valenti, S.; Mancini, A.; Giuliani, G.; Anzini, M.; Altieri, S.; Bortolussi, S.; Ferrari, C.; Clerici, A.M.; Zonta, C.; et al. Carborane-Conjugated 2-Quinolinecarboxamide Ligands of the Translocator Protein for Boron Neutron Capture Therapy. Bioconjugate Chem. 2010,21, 2213–2221. [CrossRef] 30. Worm, D.J.; Hoppenz, P.; Els-Heindl, S.; Kellert, M.; Kuhnert, R.; Saretz, S.; Köbberling, J.; Riedl, B.; Hey-Hawkins, E.; BeckSickinger, A.G. Selective Neuropeptide y Conjugates with Maximized Carborane Loading as Promising Boron Delivery Agents for Boron Neutron Capture Therapy. J. Med. Chem. 2020,63, 2358–2371. [CrossRef] 31. Feiner, I.V.J.; Pulagam, K.R.; Gómez-Vallejo, V.; Zamacola, K.; Baz, Z.; Caffarel, M.M.; Lawrie, C.H.; Ruiz-de-Angulo, A.; Carril, M.; Llop, J. Therapeutic Pretargeting with Gold Nanoparticles as Drug Candidates for Boron Neutron Capture Therapy. Part. Part. Syst. Charact. 2020,37, 2000200. [CrossRef] 32. Pulagam, K.R.; Gona, K.B.; Gómez-Vallejo, V.; Meijer, J.; Zilberfain, C.; Estrela-Lopis, I.; Baz, Z.; Cossío, U.; Llop, J. Gold Nanoparticles as Boron Carriers for Boron Neutron Capture Therapy: Synthesis, Radiolabelling and in Vivo Evaluation. Molecules 2019,24, 3609. [CrossRef] 33. Li, L.; Li, J.; Shi, Y.; Du, P.; Zhang, Z.; Liu, T.; Zhang, R.; Liu, Z. On-Demand Biodegradable Boron Nitride Nanoparticles for Treating Triple Negative Breast Cancer with Boron Neutron Capture Therapy. ACS Nano 2019 ,13, 13843–13852. [CrossRef] [PubMed] 34. Porras, I. Sulfur-33 Nanoparticles: A Monte Carlo Study of Their Potential as Neutron Capturers for Enhancing Boron Neutron Capture Therapy of Cancer. Appl. Radiat. Isot. 2011,69, 1838–1841. [CrossRef] [PubMed] 35. Barth, R.F.; Mi, P.; Yang, W. Boron Delivery Agents for Neutron Capture Therapy of Cancer. Cancer Commun. 2018 ,38, 35. [CrossRef] 36. Sauerwein, W.A.G.; Sancey, L.; Hey-Hawkins, E.; Kellert, M.; Panza, L.; Imperio, D.; Balcerzyk, M.; Rizzo, G.; Scalco, E.; Herrmann, K.; et al. Theranostics in Boron Neutron Capture Therapy. Life 2021,11, 330. [CrossRef] [PubMed] 37. Matovi´c, J.; Järvinen, J.; Sokka, I.K.; Imlimthan, S.; Raitanen, J.-E.; Montaser, A.; Maaheimo, H.; Huttunen, K.M.; Peräniemi, S.; Airaksinen, A.J.; et al. Exploring the Biochemical Foundations of a Successful GLUT1-Targeting Strategy to BNCT: Chemical Synthesis and In Vitro Evaluation of the Entire Positional Isomer Library of Ortho-Carboranylmethyl-Bearing Glucoconjugates. Mol. Pharm. 2021,18, 285–304. [CrossRef] 38. Desouky, O.; Ding, N.; Zhou, G. Targeted and Non-Targeted Effects of Ionizing Radiation. J. Radiat. Res. Appl. Sci. 2015 ,8, 247–254. [CrossRef] 39. Tian, F.; Geng, C.-R.; Tang, X.-B.; Shu, D.-Y.; Ye, H.-F.; Bortolussi, S.; Gong, C.-H. Analysis of Influencing Factors on the Method for Determining Boron Concentration and Dose through Dual Prompt Gamma Detection. Nucl. Sci. Tech. 2021 ,32, 35. [CrossRef] 40. Hu, K.; Yang, Z.; Zhang, L.; Xie, L.; Wang, L.; Xu, H.; Josephson, L.; Liang, S.H.; Zhang, M.-R. Boron Agents for Neutron Capture Therapy. Coord. Chem. Rev. 2020,405, 213139. [CrossRef] 41. Elgazzar Abdelhamid, H.; Kazem, N. Biological Effects of Ionizing Radiation. In The Pathophysiologic Basis of Nuclear Medicine; Elgazzar, A.H., Ed.; Springer: Berlin/Heidelberg, Germany, 2006; pp. 540–548, ISBN 978-3-540-47953-6. 42. Freneau, A.; dos Santos, M.; Voisin, P.; Tang, N.; Bueno Vizcarra, M.; Villagrasa, C.; Roy, L.; Vaurijoux, A.; Gruel, G. Relation between DNA Double-Strand Breaks and Energy Spectra of Secondary Electrons Produced by Different X-ray Energies. Int. J. Radiat. Biol. 2018,94, 1075–1084. [CrossRef] Cancers 2022,14, 2865 22 of 26 43. Fujita, Y.; Kato, I.; Iwai, S.; Ono, K.; Suzuki, M.; Sakurai, Y.; Ohnishi, K.; Ohnishi, T.; Yura, Y. Role of P53 Mutation in the Effect of Boron Neutron Capture Therapy on Oral Squamous Cell Carcinoma. Radiat. Oncol. 2009,4, 63. [CrossRef] 44. Takatsuji, T.; Yoshikawa, I.; Sasaki, M.S. Generalized Concept of the LET-RBE Relationship of Radiation-Induced Chromosome Aberration and Cell Death. J. Radiat. Res. 1999,40, 59–69. [CrossRef] [PubMed] 45. Maliszewska-Olejniczak, K.; Kaniowski, D.; Araszkiewicz, M.; Tymi´nska, K.; Korgul, A. Molecular Mechanisms of Specific Cellular DNA Damage Response and Repair Induced by the Mixed Radiation Field During Boron Neutron Capture Therapy. Front. Oncol. 2021,11, 1799. [CrossRef] [PubMed] 46. Galluzzi, L.; Vitale, I.; Aaronson, S.A.; Abrams, J.M.; Adam, D.; Agostinis, P.; Alnemri, E.S.; Altucci, L.; Amelio, I.; Andrews, D.W.; et al. Molecular Mechanisms of Cell Death: Recommendations of the Nomenclature Committee on Cell Death 2018. Cell Death Differ. 2018,25, 486–541. [CrossRef] 47. Wang, P.; Zhen, H.; Jiang, X.; Zhang, W.; Cheng, X.; Guo, G.; Mao, X.; Zhang, X. Boron Neutron Capture Therapy Induces Apoptosis of Glioma Cells through Bcl-2/Bax. BMC Cancer 2010,10, 661. [CrossRef] [PubMed] 48. Matsuya, Y.; Fukunaga, H.; Omura, M.; Date, H. A Model for Estimating Dose-Rate Effects on Cell-Killing of Human Melanoma after Boron Neutron Capture Therapy. Cells 2020,9, 1117. [CrossRef] 49. Seki, K.; Kinashi, Y.; Takahashi, S. Influence of P53 Status on the Effects of Boron Neutron Capture Therapy in Glioblastoma. Anticancer Res. 2015,35, 169–174. 50. Rodriguez, C.; Carpano, M.; Curotto, P.; Thorp, S.; Casal, M.; Juvenal, G.; Pisarev, M.; Dagrosa, M.A. In Vitro Studies of DNA Damage and Repair Mechanisms Induced by BNCT in a Poorly Differentiated Thyroid Carcinoma Cell Line. Radiat. Environ. Biophys. 2018,57, 143–152. [CrossRef] 51. Syamputra, D.N.I.; Sardjono, Y.; Mahmudah, R.S.N. Dose Analysis of BNCT Treatment Method for Rhabdomyosarcoma in the Head and Neck Regions Based on PHITS Code. ASEAN J. Sci. Technol. Dev. 2020,35, 235–239. [CrossRef] 52. Ono, K.; Tanaka, H.; Suzuki, M. Reevaluation of CBE Value of BPA for Hepatocytes. Appl. Radiat. Isot. 2020 ,161, 109159. [CrossRef] 53. Streitmatter, S.W.; Stewart, R.D.; Moffitt, G.; Jevremovic, T. Mechanistic Modeling of the Relative Biological Effectiveness of Boron Neutron Capture Therapy. Cells 2020,9, 2302. [CrossRef] 54. Aldossari, S.; McMahon, G.; Lockyer, N.P.; Moore, K.L. Microdistribution and Quantification of the Boron Neutron Capture Therapy Drug BPA in Primary Cell Cultures of Human Glioblastoma Tumour by NanoSIMS. Analyst 2019 ,144, 6214–6224. [CrossRef] [PubMed] 55. Swanepoel, M.W. The Role of the 14N(n,p)14C Reaction in Neutron Irradiation of Soft Tissues. Radiat. Meas. 2010 ,45, 1458–1461. [CrossRef] 56. Kumada, H.; Takada, K. Treatment Planning System and Patient Positioning for Boron Neutron Capture Therapy. Ther. Radiol. Oncol. 2018,2, 50. [CrossRef] 57. Lee, C.M.; Lee, H.S. Development of a Dose Estimation Code for BNCT with GPU Accelerated Monte Carlo and Collapsed Cone Convolution Method. Nucl. Eng. Technol. 2022,54, 1769–1780. [CrossRef] 58. Hu, N.; Tanaka, H.; Kakino, R.; Yoshikawa, S.; Miyao, M.; Akita, K.; Isohashi, K.; Aihara, T.; Nihei, K.; Ono, K. Evaluation of a Treatment Planning System Developed for Clinical Boron Neutron Capture Therapy and Validation against an Independent Monte Carlo Dose Calculation System. Radiat. Oncol. 2021,16, 243. [CrossRef] 59. Judith, F. MCNP TM-A General Monte Carlo N-Particle Transport Code Version 4C; University of California: San Diego, CA, USA, 2000. 60. Wheeler, F.J.; Wessol, D.E.; Wemple, C.A.; Nigg, D.W.; Albright, C.L.; Cohen, M.T.; Frandsen, M.W.; Harkin, G.J.; Rossmeier, M.B. SERA-an Advanced Treatment Planning System for Neutron Therapy; Idaho National Engineering and Environmental: Idaho Falls, ID, USA, 1999. 61. Iwase, H.; Niita, K.; Nakamura, T. Development of General-Purpose Particle and Heavy Ion Transport Monte Carlo Code. J. Nucl. Sci. Tehnol. 2012,39, 1142–1151. [CrossRef] 62. Hu, N.; Tanaka, H.; Takata, T.; Endo, S.; Masunaga, S.; Suzuki, M.; Sakurai, Y. Evaluation of PHITS for Microdosimetry in BNCT to Support Radiobiological Research. Appl. Radiat. Isot. 2020,161, 109148. [CrossRef] 63. Sato, T.; Masunaga, S.; Kumada, H.; Hamada, N. Microdosimetric Modeling of Biological Effectiveness for Boron Neutron Capture Therapy Considering Intraand Intercellular Heterogeneity in 10B Distribution. Sci. Rep. 2018,8, 988. [CrossRef] 64. Evaluated Nuclear Data File (ENDF). Available online: https://www.nndc.bnl.gov/exfor/endf00.jsp (accessed on 13 September 2021). 65. International Atomic Energy Agency. Current Status of Neutron Capture Therapy; IAEA: Vienna, Austria, 2001; Volume IAEATECDOC-1223. 66. Incerti, S.; Kyriakou, I.; Bernal, M.A.; Bordage, M.C.; Francis, Z.; Guatelli, S.; Ivanchenko, V.; Karamitros, M.; Lampe, N.; Lee, S.B.; et al. Geant4-DNA Example Applications for Track Structure Simulations in Liquid Water: A Report from the Geant4-DNA Project. Med. Phys. 2018,45, e722–e739. [CrossRef] 67. Perry, C.C.; Ramos-Méndez, J.; Milligan, J.R. DNA Condensation with a Boron-Containing Cationic Peptide for Modeling Boron Neutron Capture Therapy. Radiat. Phys. Chem. 2020,166, 108521. [CrossRef] 68. Sun, T.; Zhang, Z.; Li, B.; Chen, G.; Xie, X.; Wei, Y.; Wu, J.; Zhou, Y.; Du, Z. Boron Neutron Capture Therapy Induces Cell Cycle Arrest and Cell Apoptosis of Glioma Stem/Progenitor Cells in Vitro. Radiat. Oncol. 2013,8, 195. [CrossRef] Cancers 2022,14, 2865 23 of 26 69. Pedrosa-Rivera, M.; Praena, J.; Porras, I.; Sabariego, M.P.; Köster, U.; Haertlein, M.; Forsyth, V.T.; Ramírez, J.C.; Jover, C.; Jimena, D.; et al. Thermal Neutron Relative Biological Effectiveness Factors for Boron Neutron Capture Therapy from In Vitro Irradiations. Cells 2020,9, 2144. [CrossRef] [PubMed] 70. Staaf, E.; Brehwens, K.; Haghdoost, S.; Czub, J.; Wojcik, A. Gamma-H2AX Foci in Cells Exposed to a Mixed Beam of X-rays and Alpha Particles. Genome Integr. 2012,3, 8. [CrossRef] [PubMed] 71. Masutani, M.; Baiseitov, D.; Itoh, T.; Hirai, T.; Berikkhanova, K.; Murakami, Y.; Zhumadilov, Z.; Imahori, Y.; Hoshi, M.; Itami, J. Histological and Biochemical Analysis of DNA Damage after BNCT in Rat Model. Appl. Radiat. Isot. 2014 ,88, 104–108. [CrossRef] [PubMed] 72. Lapytsko, A.; Kollarovic, G.; Ivanova, L.; Studencka, M.; Schaber, J. FoCo: A Simple and Robust Quantification Algorithm of Nuclear Foci. BMC Bioinform. 2015,16, 392. [CrossRef] [PubMed] 73. López-Valverde, J.A.; Jiménez-Ortega, E.; Leal, A. Clinical Feasibility Study of Gold Nanoparticles as Theragnostic Agents for Precision Radiotherapy. Biomedicines 2022,10, 1214. [CrossRef] 74. Kuperman, V.Y. Effect of Dose Rate in Hypofractionated Radiotherapy. Phys. Med. 2019,65, 191–199. [CrossRef] 75. Faião-Flores, F.; Coelho, P.R.P.; Muniz, R.O.R.; Souza, G.S.; Arruda-Neto, J.; Maria, D.A. Antitumor Potential Induction and Free Radicals Production in Melanoma Cells by Boron Neutron Capture Therapy. Appl. Radiat. Isot. 2011,69, 1748–1751. [CrossRef] 76. Mares, V.; Krajcí, D.; Lisá, V. Subcellular Targets of Mercaptoborate (BSH), a Carrier of 10B for Neutron Capture Therapy (BNCT) of Brain Tumors. Physiol. Res. 2003,52, 629–636. 77. Romanov, V.; Isohashi, K.; Alobthani, G.; Beshr, R.; Horitsugi, G.; Kanai, Y.; Naka, S.; Watabe, T.; Shimosegawa, E.; Hatazawa, J. Evaluation of the Total Distribution Volume of 18F-FBPA in Normal Tissues of Healthy Volunteers by Non-Compartmental Kinetic Modeling. Ann. Nucl. Med. 2020,34, 155–162. [CrossRef] 78. Coderre, J.A.; Chanana, A.D.; Joel, D.D.; Elowitz, E.H.; Micca, P.L.; Nawrocky, M.M.; Chadha, M.; Gebbers, J.O.; Shady, M.; Peress, N.S.; et al. Biodistribution of Boronophenylalanine in Patients with Glioblastoma Multiforme: Boron Concentration Correlates with Tumor Cellularity. Radiat. Res. 1998,149, 163–170. [CrossRef] [PubMed] 79. Fukumitsu, N.; Matsumoto, Y. Development of an Imaging Technique for Boron Neutron Capture Therapy. Cells 2021 ,10, 2135. [CrossRef] [PubMed] 80. Menichetti, L.; Petroni, D.; Panetta, D.; Burchielli, S.; Bortolussi, S.; Matteucci, M.; Pascali, G.; del Turco, S.; del Guerra, A.; Altieri, S.; et al. A Micro-PET/CT Approach Using O-(2-[18F]Fluoroethyl)-l-Tyrosine in an Experimental Animal Model of F98 Glioma for BNCT. Appl. Radiat. Isot. 2011,69, 1717–1720. [CrossRef] [PubMed] 81. Shimosegawa, E.; Isohashi, K.; Naka, S.; Horitsugi, G.; Hatazawa, J. Assessment of 10B Concentration in Boron Neutron Capture Therapy: Potential of Image-Guided Therapy Using 18FBPA PET. Ann. Nucl. Med. 2016,30, 749–755. [CrossRef] [PubMed] 82. Li, J.; Shi, Y.; Zhang, Z.; Liu, H.; Lang, L.; Liu, T.; Chen, X.; Liu, Z. A Metabolically Stable Boron-Derived Tyrosine Serves as a Theranostic Agent for Positron Emission Tomography Guided Boron Neutron Capture Therapy. Bioconjugate Chem. 2019 ,30, 2870–2878. [CrossRef] 83. Sato, A.; Itoh, T.; Imamichi, S.; Kikuhara, S.; Fujimori, H.; Hirai, T.; Saito, S.; Sakurai, Y.; Tanaka, H.; Nakamura, H.; et al. Proteomic Analysis of Cellular Response Induced by Boron Neutron Capture Reaction in Human Squamous Cell Carcinoma SAS Cells. Appl. Radiat. Isot. 2015,106, 213–219. [CrossRef] [PubMed] 84. Ferrari, E.; Wittig, A.; Basilico, F.; Rossi, R.; de Palma, A.; di Silvestre, D.; Sauerwein, W.; Mauri, P. Urinary Proteomics Profiles Are Useful for Detection of Cancer Biomarkers and Changes Induced by Therapeutic Procedures. Molecules 2019 ,24, 794. [CrossRef] 85. Ekholm, F.S.; Matovic, J.; Jarvinen, J.; Bland, H.C.; Sokka, I.K.; Imlimthan, S.; Huttunen, K.M.; Timonen, J.; Peraniemi, S.; Aitio, O.; et al. Addressing the Biochemical Foundations of a Glucose-Based “Trojan Horse”-Strategy to Boron Neutron Capture Therapy: From Chemical Synthesis to in Vitro Assessment. Mol. Pharm. 2020,17, 3885–3899. [CrossRef] 86. Balcerzyk, M.; De-Miguel, M.; Guerrero, C.; Fernandez, B. Quantification of Boron Compound Concentration for BNCT Using Positron Emission Tomography. Cells 2020,9, 2084. [CrossRef] 87. Nariai, T.; Ishiwata, K.; Kimura, Y.; Inaji, M.; Momose, T.; Yamamoto, T.; Matsumura, A.; Ishii, K.; Ohno, K. PET Pharmacokinetic Analysis to Estimate Boron Concentration in Tumor and Brain as a Guide to Plan BNCT for Malignant Cerebral Glioma. Appl. Radiat. Isot. 2009,67, S348–S350. [CrossRef] 88. Watanabe, Y.; Kurihara, H.; Itami, J.; Sasaki, R.; Arai, Y.; Sugimura, K. Relationship between the Uptake of 18F-Borono-LPhenylalanine and L-[Methyl-11C] Methionine in Head and Neck Tumors and Normal Organs. Radiat. Oncol. 2017 ,12, 17. [CrossRef] [PubMed] 89. Yamamoto, T.; Nakai, K.; Nariai, T.; Kumada, H.; Okumura, T.; Mizumoto, M.; Tsuboi, K.; Zaboronok, A.; Ishikawa, E.; Aiyama, H.; et al. The Status of Tsukuba BNCT Trial: BPA-Based Boron Neutron Capture Therapy Combined with X-ray Irradiation. Appl. Radiat. Isot. 2011,69, 1817–1818. [CrossRef] [PubMed] 90. Alberti, D.; Deagostino, A.; Toppino, A.; Protti, N.; Bortolussi, S.; Altieri, S.; Aime, S.; Geninatti Crich, S. An Innovative Therapeutic Approach for Malignant Mesothelioma Treatment Based on the Use of Gd/Boron Multimodal Probes for MRI Guided BNCT. J. Control. Release 2018,280, 31–38. [CrossRef] 91. Geninatti-Crich, S.; Deagostino, A.; Toppino, A.; Alberti, D.; Venturello, P.; Aime, S. Boronated Compounds for Imaging Guided BNCT Applications. Anti-Cancer Agents Med. Chem. 2012,12, 543–553. [CrossRef] Cancers 2022,14, 2865 24 of 26 92. Geninatti-Crich, S.; Alberti, D.; Szabo, I.; Deagostino, A.; Toppino, A.; Barge, A.; Ballarini, F.; Bortolussi, S.; Bruschi, P.; Protti, N.; et al. MRI-Guided Neutron Capture Therapy by Use of a Dual Gadolinium/Boron Agent Targeted at Tumour Cells through Upregulated Low-Density Lipoprotein Transporters. Chem. Eur. J. 2011,17, 8479–8486. [CrossRef] [PubMed] 93. Wu, C.-Y.; Hsieh, H.-H.; Chang, T.-Y.; Lin, J.-J.; Wu, C.-C.; Hsu, M.-H.; Lin, M.-C.; Peng, S.-L. Development of MRI-Detectable Boron-Containing Gold Nanoparticle-Encapsulated Biodegradable Polymeric Matrix for Boron Neutron Capture Therapy (BNCT). Int. J. Mol. Sci. 2021,22, 8050. [CrossRef] [PubMed] 94. Kuthala, N.; Vankayala, R.; Li, Y.; Chiang, C.; Hwang, K.C. Engineering Novel Targeted Boron-10-Enriched Theranostic Nanomedicine to Combat against Murine Brain Tumors via MR Imaging-Guided Boron Neutron Capture Therapy. Adv. Mater. 2017,29, 1700850. [CrossRef] 95. Bonora, M.; Corti, M.; Borsa, F.; Bortolussi, S.; Protti, N.; Santoro, D.; Stella, S.; Altieri, S.; Zonta, C.; Clerici, A.M.; et al. 1H and 10B NMR and MRI Investigation of Boronand Gadolinium–Boron Compounds in Boron Neutron Capture Therapy. Appl. Radiat. Isot. 2011,69, 1702–1705. [CrossRef] 96. Chandra, S.; Ahamad, T.; Barth, R.F.; Kabalka, G.W. Quantitative Evaluation of Boron Neutron Capture Therapy (BNCT) Drugs for Boron Delivery and Retention at Subcellular-Scale Resolution in Human Glioblastoma Cells with Imaging Secondary Ion Mass Spectrometry (SIMS). J. Microsc. 2014,254, 146–156. [CrossRef] 97. Lanni, E.J.; Rubakhin, S.S.; Sweedler, J.V. Mass Spectrometry Imaging and Profiling of Single Cells. J. Proteom. 2012 ,75, 5036–5051. [CrossRef] 98. Barth, R.F.; Kabalka, G.W.; Yang, W.; Huo, T.; Nakkula, R.J.; Shaikh, A.L.; Haider, S.A.; Chandra, S. Evaluation of Unnatural Cyclic Amino Acids as Boron Delivery Agents for Treatment of Melanomas and Gliomas. Appl. Radiat. Isot. 2014 ,88, 38–42. [CrossRef] [PubMed] 99. Arlinghaus, H.F.; Fartmann, M.; Kriegeskotte, C.; Dambach, S.; Wittig, A.; Sauerwein, W.; Lipinsky, D. Subcellular Imaging of Cell Cultures and Tissue for Boron Localization with Laser-SNMS. Surf. Interface Anal. 2004,36, 698–701. [CrossRef] 100. Chandra, S.; Barth, R.F.; Haider, S.A.; Yang, W.; Huo, T.; Shaikh, A.L.; Kabalka, G.W. Biodistribution and Subcellular Localization of an Unnatural Boron-Containing Amino Acid (Cis-ABCPC) by Imaging Secondary Ion Mass Spectrometry for Neutron Capture Therapy of Melanomas and Gliomas. PLoS ONE 2013,8, e75377. [CrossRef] [PubMed] 101. Murata, I.; Kusaka, S.; Minami, K.; Saraue, N.; Tamaki, S.; Kato, I.; Sato, F. Design of SPECT for BNCT to Measure Local Boron Dose with GAGG Scintillator. Appl. Radiat. Isot. 2022,181, 110056. [CrossRef] [PubMed] 102. Minsky, D.M.; Valda, A.A.; Kreiner, A.J.; Green, S.; Wojnecki, C.; Ghani, Z. First Tomographic Image of Neutron Capture Rate in a BNCT Facility. Appl. Radiat. Isot. 2011,69, 1858–1861. [CrossRef] [PubMed] 103. Minsky, D.M.; Valda, A.A.; Kreiner, A.J.; Green, S.; Wojnecki, C.; Ghani, Z. Experimental Feasibility Studies on a SPECT Tomograph for BNCT Dosimetry. Appl. Radiat. Isot. 2009,67, S179–S182. [CrossRef] 104. Winkler, A.; Koivunoro, H.; Reijonen, V.; Auterinen, I.; Savolainen, S. Prompt Gamma and Neutron Detection in BNCT Utilizing a CdTe Detector. Appl. Radiat. Isot. 2015,106, 139–144. [CrossRef] 105. Abbene, L.; Principato, F.; Buttacavoli, A.; Gerardi, G.; Bettelli, M.; Zappettini, A.; Altieri, S.; Auricchio, N.; Caroli, E.; Zanettini, S.; et al. Potentialities of High-Resolution 3-D CZT Drift Strip Detectors for Prompt Gamma-Ray Measurements in BNCT. Sensors 2022,22, 1502. [CrossRef] 106. Murata, I.; Mukai, T.; Nakamura, S.; Miyamaru, H.; Kato, I. Development of a Thick CdTe Detector for BNCT–SPECT. Appl. Radiat. Isot. 2011,69, 1706–1709. [CrossRef] 107. Murata, I.; Nakamura, S.; Manabe, M.; Miyamaru, H.; Kato, I. Characterization Measurement of a Thick CdTe Detector for BNCT-SPECT—Detection Efficiency and Energy Resolution. Appl. Radiat. Isot. 2014,88, 129–133. [CrossRef] 108. Kim, M.; Hong, B.H.; Cho, I.; Park, C.; Min, S.-H.; Hwang, W.T.; Lee, W.; Kim, K.M. Design of a Scintillator-Based Prompt Gamma Camera for Boron-Neutron Capture Therapy: Comparison of SrI2 and GAGG Using Monte-Carlo Simulation. Nucl. Eng. Technol. 2021,53, 626–636. [CrossRef] 109. Kanno, I.; Nishimatsu, D.; Funama, F. Simulation Study on the Feasibility of Current-Mode SPECT for B-10 Concentration Estimation in Boron Neutron Capture Therapy. J. Instrum. 2019,14, C02002. [CrossRef] 110. Goodman, J.H.; Yang, W.; Barth, R.F.; Gao, Z.; Boesel, C.P.; Staubus, A.E.; Gupta, N.; Gahbauer, R.A.; Adams, D.M.; Gibson, C.R.; et al. Boron Neutron Capture Therapy of Brain Tumors: Biodistribution, Pharmacokinetics, and Radiation Dosimetry Sodium Borocaptate in Patients with Gliomas. Neurosurgery 2000,47, 608–622. [CrossRef] [PubMed] 111. Verbakel, W.F.A.R. Validation of the Scanning γ -Ray Telescope for in Vivo Dosimetry and Boron Measurements during BNCT. Phys. Med. Biol. 2001,46, 3269. [CrossRef] [PubMed] 112. Moro, D.; Colautti, P.; Gualdrini, G.; Masi, M.; Conte, V.; de Nardo, L.; Tornielli, G. Two Miniaturised TEPCS in a Single Detector for BNCT Microdosimetry. Radiat. Prot. Dosim. 2006,122, 396–400. [CrossRef] [PubMed] 113. Moro, D.; Colautti, P.; Lollo, M.; Esposito, J.; Conte, V.; de Nardo, L.; Ferretti, A.; Ceballos, C. BNCT Dosimetry Performed with a Mini Twin Tissue-Equivalent Proportional Counters (TEPC). Appl. Radiat. Isot. 2009,67, S171–S174. [CrossRef] [PubMed] 114. Kiyanagi, Y.; Sakurai, Y.; Kumada, H.; Tanaka, H. Status of Accelerator-Based BNCT Projects Worldwide. AIP Conf. Proc. 2019 , 2160, 050012. 115. Kato, T.; Hirose, K.; Tanaka, H.; Mitsumoto, T.; Motoyanagi, T.; Arai, K.; Harada, T.; Takeuchi, A.; Kato, R.; Yajima, S.; et al. Design and Construction of an Accelerator-Based Boron Neutron Capture Therapy (AB-BNCT) Facility with Multiple Treatment Rooms at the Southern Tohoku BNCT Research Center. Appl. Radiat. Isot. 2020,156, 108961. [CrossRef] Cancers 2022,14, 2865 25 of 26 116. BNCT System NeuCure ® |BNCT System MEDICAL|Sumitomo Heavy Industries, Ltd. Industrial Equipment Division. Available online: https://www.shi.co.jp/industrial/en/product/medical/bnct/neucure.html (accessed on 30 April 2022). 117. IBA Worldwide—Shaping the Future of Proton Therapy. Available online: https://www.iba-worldwide.com/proton-therapy (accessed on 30 April 2022). 118. He, H.; Li, J.; Jiang, P.; Tian, S.; Wang, H.; Fan, R.; Liu, J.; Yang, Y.; Liu, Z.; Wang, J. The Basis and Advances in Clinical Application of Boron Neutron Capture Therapy. Radiat. Oncol. 2021,16, 216. [CrossRef] 119. Asbury, A.K.; Ojemann, R.G.; Nielsen, S.L.; Sweet, W.H. Neuropathologic Study of Fourteen Cases of Malignant Brain Tumor Treated by Boron-10 Slow Neutron Capture Radiation. J. Neuropathol. Exp. Neurol. 1972,31, 278–303. [CrossRef] 120. Sweet, W.H.; Javid, M. The Possible Use of Neutron-Capturing Isotopes Such as Boron 10 in the Treatment of Neoplasms. I. Intracranial Tumors. J. Neurosurg. 1952,9, 200–209. [CrossRef] [PubMed] 121. Mishima, Y.; Ichihashi, M.; Hatta, S.; Honda, C.; Yamamura, K.; Nakagawa, T.; Obara, H.; Shirakawa, J.; Hiratsuka, J.; Taniyama, K. First Human Clinical Trial of Melanoma Neutron Capture. Diagnosis and Therapy. Strahlenther. Onkol. 1989 ,165, 251–254. [PubMed] 122. Joensuu, H.; Kankaanranta, L.; Seppälä, T.; Auterinen, I.; Kallio, M.; Kulvik, M.; Laakso, J.; Vähätalo, J.; Kortesniemi, M.; Kotiluoto, P.; et al. Boron Neutron Capture Therapy of Brain Tumors: Clinical Trials at the Finnish Facility Using Boronophenylalanine. J. Neuro-Oncol. 2003,62, 123–134. [CrossRef] 123. Kankaanranta, L.; Seppälä, T.; Koivunoro, H.; Saarilahti, K.; Atula, T.; Collan, J.; Salli, E.; Kortesniemi, M.; Uusi-Simola, J.; Mäkitie, A.; et al. Boron Neutron Capture Therapy in the Treatment of Locally Recurred Head and Neck Cancer. Int. J. Radiat. Oncol. Biol. Phys. 2007,69, 475–482. [CrossRef] [PubMed] 124. Kankaanranta, L.; Seppälä, T.; Koivunoro, H.; Saarilahti, K.; Atula, T.; Collan, J.; Salli, E.; Kortesniemi, M.; Uusi-Simola, J.; Välimäki, P.; et al. Boron Neutron Capture Therapy in the Treatment of Locally Recurred Head-and-Neck Cancer: Final Analysis of a Phase I/II Trial. Int. J. Radiat. Oncol. Biol. Phys. 2012,82, e67–e75. [CrossRef] 125. Kankaanranta, L.; Seppälä, T.; Koivunoro, H.; Välimäki, P.; Beule, A.; Collan, J.; Kortesniemi, M.; Uusi-Simola, J.; Kotiluoto, P.; Auterinen, I.; et al. L-Boronophenylalanine-Mediated Boron Neutron Capture Therapy for Malignant Glioma Progressing after External Beam Radiation Therapy: A Phase i Study. Int. J. Radiat. Oncol. Biol. Phys. 2011,80, 369–376. [CrossRef] 126. Koivunoro, H.; Hippeläinen, E.; Auterinen, I.; Kankaanranta, L.; Kulvik, M.; Laakso, J.; Seppälä, T.; Savolainen, S.; Joensuu, H. Biokinetic Analysis of Tissue Boron (10B) Concentrations of Glioma Patients Treated with BNCT in Finland. Appl. Radiat. Isot. 2015,106, 189–194. [CrossRef] [PubMed] 127. Cruickshank, G.S.; Ngoga, D.; Detta, A.; Green, S.; James, N.D.; Wojnecki, C.; Doran, J.; Hardie, J.; Chester, M.; Graham, N.; et al. A Cancer Research UK Pharmacokinetic Study of BPA-Mannitol in Patients with High Grade Glioma to Optimise Uptake Parameters for Clinical Trials of BNCT. Appl. Radiat. Isot. 2009,67, S31–S33. [CrossRef] 128. Yong, Z.; Song, Z.; Zhou, Y.; Liu, T.; Zhang, Z.; Zhao, Y.; Chen, Y.; Jin, C.; Chen, X.; Lu, J.; et al. Boron Neutron Capture Therapy for Malignant Melanoma: First Clinical Case Report in China. Chin. J. Cancer Res. 2016,28, 634. [CrossRef] 129. Wang, L.-W.; Chen, Y.-W.; Ho, C.-Y.; Hsueh Liu, Y.-W.; Chou, F.-I.; Liu, Y.-H.; Liu, H.-M.; Peir, J.-J.; Jiang, S.-H.; Chang, C.-W.; et al. Fractionated Boron Neutron Capture Therapy in Locally Recurrent Head and Neck Cancer: A Prospective Phase I/II Trial. Int. J. Radiat. Oncol. Biol. Phys. 2016,95, 396–403. [CrossRef] 130. Wang, L.W.; Liu, Y.W.H.; Chou, F.I.; Jiang, S.H. Clinical Trials for Treating Recurrent Head and Neck Cancer with Boron Neutron Capture Therapy Using the Tsing-Hua Open Pool Reactor. Cancer Commun. 2018,38, 37. [CrossRef] [PubMed] 131. Lee, J.-C.; Chuang, K.-S.; Hsueh Liu, Y.-W.; Lin, T.-Y.; Teng, Y.-C.; Wang, L.-W. A Comparison of Dose Distributions in Gross Tumor Volume between Boron Neutron Capture Therapy Alone and Combined Boron Neutron Capture Therapy plus Intensity Modulation Radiation Therapy for Head and Neck Cancer. PLoS ONE 2019,14, e0210626. [CrossRef] 132. Lan, T.-L.; Chou, F.-I.; Lin, K.-H.; Pan, P.-S.; Lee, J.-C.; Huang, W.-S.; Liu, Y.-M.; Chao, Y.; Chen, Y.-W. Using Salvage Boron Neutron Capture Therapy (BNCT) for Recurrent Malignant Brain Tumors in Taiwan. Appl. Radiat. Isot. 2020 ,160, 109105. [CrossRef] [PubMed] 133. Chen, Y.-W.; Lee, Y.-Y.; Lin, C.-F.; Pan, P.-S.; Chen, J.-K.; Wang, C.-W.; Hsu, S.-M.; Kuo, Y.-C.; Lan, T.-L.; Hsu, S.P.C.; et al. Salvage Boron Neutron Capture Therapy for Malignant Brain Tumor Patients in Compliance with Emergency and Compassionate Use: Evaluation of 34 Cases in Taiwan. Biology 2021,10, 334. [CrossRef] 134. Ree, A.H.; Redalen, K.R. Personalized Radiotherapy: Concepts, Biomarkers and Trial Design. Br. J. Radiol. 2015 ,88, 20150009. [CrossRef] [PubMed] 135. Formenti, S.C.; Demaria, S. Combining Radiotherapy and Cancer Immunotherapy: A Paradigm Shift. J. Natl. Cancer Inst. 2013 , 105, 256–265. [CrossRef] [PubMed] 136. Giaccia, A.J. Molecular Radiobiology: The State of the Art. J. Clin. Oncol. 2014,32, 2871–2878. [CrossRef] [PubMed] 137. Sanduleanu, S.; van der Wiel, A.M.A.; Lieverse, R.I.Y.; Marcus, D.; Ibrahim, A.; Primakov, S.; Wu, G.; Theys, J.; Yaromina, A.; Dubois, L.J.; et al. Hypoxia PET Imaging with [18F]-HX4—A Promising Next-Generation Tracer. Cancers 2020 ,12, 1322. [CrossRef] 138. McKinley, E.T.; Ayers, G.D.; Smith, R.A.; Saleh, S.A.; Zhao, P.; Washington, M.K.; Coffey, R.J.; Manning, H.C. Limits of [18F]-FLT PET as a Biomarker of Proliferation in Oncology. PLoS ONE 2013,8, e58938. [CrossRef] 139. Sharma, S. PET Radiopharmaceuticals for Personalized Medicine. Curr. Drug Targets 2016,17, 1894–1907. [CrossRef] 140. Jiménez-Ortega, E.; Ureba, A.; Vargas, A.; Baeza, J.A.; Wals-Zurita, A.; García-Gómez, F.J.; Barbeiro, A.R.; Leal, A. Dose Painting by Means of Monte Carlo Treatment Planning at the Voxel Level. Phys. Med. 2017,42, 339–344. [CrossRef] [PubMed]