Encapsulating Nickel-Iron Alloy Nanoparticles in a Polysilazane-Derived Microporous Si-C-O-N-Based Support to Stimulate Superior OER Activity.

The in situ confinement of nickel (Ni)-iron (Fe) nanoparticles (NPs) in a polymer-derived microporous silicon carboxynitride (Si-C-O-N)-based support is investigated to stimulate superior oxygen evolution reaction (OER) activity in an alkaline media. Firstly, we consider a commercial polysilazane (PSZ) and Ni and Fe chlorides to be mixed in N,N-dimethylformamide (DMF) and deliver after overnight solvent reflux a series of Ni-Fe : organosilicon coordination polymers. The latter are then heat-treated at 500 °C in flowing argon to form the title compounds. By considering a Ni : Fe ratio of 1.5, face centred cubic (fcc) NixFey alloy NPs with a size of 15-30 nm are in situ generated in a porous Si-C-O-N-based matrix displaying a specific surface area (SSA) as high as 237 m2 ⋅ g-1. Hence, encapsulated NPs are rendered accessible to promote electrocatalytic water oxidation. An OER overpotential as low as 315 mV at 10 mA ⋅ cm-2 is measured. This high catalytic performance (considering that the metal mass loading is as low as 0.24 mg cm-2) is rather stable as observed after an activation step; thus, validating our synthesis approach. This is clearly attributed to both the strong NP-matrix interaction and the confinement effect of the matrix as highlighted through post mortem microscopy observations.

Application of Cleavable Linkers to Improve Therapeutic Index of Radioligand Therapies.

Radioligand therapy (RLT) is an emergent drug class for cancer treatment. The dose administered to cancer patients is constrained by the radiation exposure to normal tissues to maintain an appropriate therapeutic index. When a radiopharmaceutical or its radiometabolite is retained in the kidneys, radiation dose deposition in the kidneys can become a dose-limiting factor. A good exemplar is [177Lu]Lu-DOTATATE, where patients receive a co-infusion of basic amino acids for nephroprotection. Besides peptides, there are other classes of targeting vectors like antibody fragments, antibody mimetics, peptidomimetics, and small molecules that clear through the renal pathway. In this review, we will review established and emerging strategies that can be used to mitigate radiation-induced nephrotoxicity, with a focus on the development and incorporation of cleavable linkers for radiopharmaceutical designs. Finally, we offer our perspectives on cleavable linkers for RLT, highlighting future areas of research that will help advance the technology.

Trastuzumab-conjugated oxine-based ligand for [89Zr]Zr4+ immunoPET.

A new, bifunctional chelating ligand for immuno-Positron Emission Tomography (PET) was designed, synthesized, and conjugated to Trastuzumab for a proof-of-concept study with 89Zr. H4neunox was synthesized from the tris(2-aminoethyl)amine backbone, decorated with 8-hydroxyquinoline moieties, and utilizes a primary amine for functionalization. A maleimide moiety extends the chelator to create H4neunox-mal for antibody conjugation via maleimide-thiol click chemistry. Preliminary 89Zr radiolabeling of H4neunox indicated quantitative radiolabeling at 1 × 10-5 M, but improved inertness towards human serum (96% intact at 7 d) and Fe3+ (92% intact at 24 h) compared to the previously synthesized H5decaox. The chelator was successfully conjugated to the monoclonal antibody, Trastuzumab, and used in preliminary radiolabeling reactions (37 °C, 2 h) with 89Zr. Radiochemical assessments of the new H4neunox-Trastuzumab conjugate include 89Zr radiolabeling, spin filter purification, cell-binding immunoreactivity, and in vivo PET imaging and biodistribution in SKOV-3 tumour bearing nude mice, performed in comparison with the desferrioxamine B analog, DFO-Trastuzumab. The [89Zr]Zr(neunox-Trastuzumab) showed lowered inertness towards serum (76% intact at 24 h) as well as demetallation in vivo through bone uptake (21% ID/g) in PET imaging and biodistribution studies when compared to [89Zr]Zr(DFO-Trastuzumab). Although the combination of the chelator and antibody had detrimental effects on their intended purposes, nonetheless, the primary amine platform of H4neunox developed here provides an oxine-based bifunctional ligand for further derivatizations with other targeting vectors.

Development of a multi faceted platform containing a tetrazine, fluorophore and chelator: synthesis, characterization, radiolabeling, and immuno-SPECT imaging.

BACKGROUND: Combining optical (fluorescence) imaging with nuclear imaging has the potential to offer a powerful tool in personal health care, where nuclear imaging offers in vivo functional whole-body visualization, and the fluorescence modality may be used for image-guided tumor resection. Varying chemical strategies have been exploited to fuse both modalities into one molecular entity. When radiometals are employed in nuclear imaging, a chelator is typically inserted into the molecule to facilitate radiolabeling; the availability of the chelator further expands the potential use of these platforms for targeted radionuclide therapy if a therapeutic radiometal is employed. Herein, a novel mixed modality scaffold which contains a tetrazine (Tz)--for biomolecule conjugation, fluorophore-for optical imaging, and chelator-for radiometal incorporation, in one construct is presented. The novel platform was characterized for its fluorescence properties, radiolabeled with single-photon emission computed tomography (SPECT) isotope indium-111 (111In3+) and therapeutic alpha emitter actinium-225 (225Ac3+). Both radiolabels were conjugated in vitro to trans-cyclooctene (TCO)-modified trastuzumab; biodistribution and immuno-SPECT imaging of the former conjugate was assessed. RESULTS: Key to the success of the platform synthesis was incorporation of a 4,4'-dicyano-BODIPY fluorophore. The route gives access to an advanced intermediate where final chelator-incorporated compounds can be easily accessed in one step prior to radiolabeling or biomolecule conjugation. The DOTA (1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid) conjugate was prepared, displayed good fluorescence properties, and was successfully radiolabeled with 111In & 225Ac in high radiochemical yield. Both complexes were then separately conjugated in vitro to TCO modified trastuzumab through an inverse electron demand Diels-Alder (IEDDA) reaction with the Tz. Pilot small animal in vivo immuno-SPECT imaging with [111In]In-DO3A-BODIPY-Tz-TCO-trastuzumab was also conducted and exhibited high tumor uptake (21.2 ± 5.6%ID/g 6 days post-injection) with low uptake in non-target tissues. CONCLUSIONS: The novel platform shows promise as a multi-modal probe for theranostic applications. In particular, access to an advanced synthetic intermediate where tailored chelators can be incorporated in the last step of synthesis expands the potential use of the scaffold to other radiometals. Future studies including validation of ex vivo fluorescence imaging and exploiting the pre-targeting approach available through the IEDDA reaction are warranted.

Implications of physics, chemistry and biology for dosimetry calculations using theranostic pairs.

Theranostics is an emerging paradigm that combines imaging and therapy in order to personalize patient treatment. In nuclear medicine, this is achieved by using radiopharmaceuticals that target identical molecular targets for both imaging (using emitted gamma rays) and radiopharmaceutical therapy (using emitted beta, alpha or Auger-electron particles) for the treatment of various diseases, such as cancer. If the therapeutic radiopharmaceutical cannot be imaged quantitatively, a "theranostic pair" imaging surrogate can be used to predict the absorbed radiation doses from the therapeutic radiopharmaceutical. However, theranostic dosimetry assumes that the pharmacokinetics and biodistributions of both radiopharmaceuticals in the pair are identical or very similar, an assumption that still requires further validation for many theranostic pairs. In this review, we consider both same-element and different-element theranostic pairs and attempt to determine if factors exist which may cause inaccurate dose extrapolations in theranostic dosimetry, either intrinsic (e.g. chemical differences) or extrinsic (e.g. injecting different amounts of each radiopharmaceutical) to the radiopharmaceuticals. We discuss the basis behind theranostic dosimetry and present common theranostic pairs and their therapeutic applications in oncology. We investigate general factors that could create alterations in the behavior of the radiopharmaceuticals or the quantitative accuracy of imaging them. Finally, we attempt to determine if there is evidence showing some specific pairs as suitable for theranostic dosimetry. We show that there are a variety of intrinsic and extrinsic factors which can significantly alter the behavior among pairs of radiopharmaceuticals, even if they belong to the same chemical element. More research is needed to determine the impact of these factors on theranostic dosimetry estimates and on patient outcomes, and how to correctly account for them.

Targeting Refractory Mantle Cell Lymphoma for Imaging and Therapy Using C-X-C Chemokine Receptor Type 4 Radioligands.

PURPOSE: Mantle cell lymphoma (MCL) is associated with poor survival. The purpose of this study was to assess whether the C-X-C chemokine receptor type 4 (CXCR4) is a useful target for imaging and radioligand therapy of MCL, using a novel pair of radioligands, [68Ga]Ga and [177Lu]Lu-BL02. EXPERIMENTAL DESIGN: We performed a retrospective analysis of 146 patients with MCL to evaluate CXCR4 expression and its correlation with outcomes. Guided by in silico methods, we designed BL02, a new radioligand labelled with 68Ga or 177Lu for PET imaging and therapy, respectively. We performed imaging and biodistribution studies in xenograft models with varying CXCR4 expression. We evaluated [177Lu]Lu-BL02 in MCL models, and evaluated its potential for therapy in Z138 MCL xenografts. RESULTS: Phosphorylated and nonphosphorylated CXCR4 expression were correlated with poor survival in patients with MCL and characterized by unique underlying molecular signatures. [68Ga]Ga-BL02 uptake correlated with CXCR4 expression, and localized lesions in a metastatic xenograft model. [177Lu]Lu-BL02 showed high uptake in MCL xenografts. Therapy studies with a single dose in the Z138 model showed tumor regression and improved survival compared with a control group. Upon regrowth, the treated mice experienced concurrent metastasis alongside localized xenograft regrowth, and recurrent lesions showed enhanced CXCR4 signaling. CONCLUSIONS: CXCR4 is an independent factor of poor prognosis for MCL and a promising target for imaging and radioligand therapy. [68Ga]Ga-BL02 showed high contrast to visualize CXCR4-expressing xenografts for PET imaging and [177Lu]Lu-BL02 induced rapid tumor regression in a preclinical model of MCL.

225Ac-H4py4pa for Targeted Alpha Therapy.

Herein, we present the syntheses and characterization of a new undecadendate chelator, H4py4pa, and its bifunctional analog H4py4pa-phenyl-NCS, conjugated to the monoclonal antibody, Trastuzumab, which targets the HER2+ cancer. H4py4pa possesses excellent affinity for 225Ac (α, t1/2 = 9.92 d) for targeted alpha therapy (TAT), where quantitative radiolabeling yield was achieved at ambient temperature, pH = 7, in 30 min at 10-6 M chelator concentration, leading to a complex highly stable in mouse serum for at least 9 d. To investigate the chelation of H4py4pa with large metal ions, lanthanum (La3+), which is the largest nonradioactive metal of the lanthanide series, was adopted as a surrogate for 225Ac to enable a series of nonradioactive chemical studies. In line with the 1H NMR spectrum, the DFT (density functional theory)-calculated structure of the [La(py4pa)]- anion possessed a high degree of symmetry, and the La3+ ion was secured by two distinct pairs of picolinate arms. Furthermore, the [La(py4pa)]- complex also demonstrated a superb thermodynamic stability (log K[La(py4pa)]- ∼ 20.33, pLa = 21.0) compared to those of DOTA (log K[La(DOTA)]- ∼ 24.25, pLa = 19.2) or H2macropa (log K[La(macropa)]- = 14.99, pLa ∼ 8.5). Moreover, the functional versatility offered by the bifunctional py4pa precursor permits facile incorporation of various linkers for bioconjugation through direct nucleophilic substitution. In this work, a short phenyl-NCS linker was incorporated to tether H4py4pa to Trastuzumab. Radiolabeling studies, in vitro serum stability, and animal studies were performed in parallel with the DOTA-benzyl-Trastuzumab. Both displayed excellent in vivo stability and tumor specificity.

A Systematic Review of Molecular Imaging Agents Targeting Bradykinin B1 and B2 Receptors.

Kinins, bradykinin and kallidin are vasoactive peptides that signal through the bradykinin B1 and B2 receptors (B1R and B2R). B2R is constitutively expressed in healthy tissues and mediates responses such as vasodilation, fluid balance and retention, smooth muscle contraction, and algesia, while B1R is absent in normal tissues and is induced by tissue trauma or inflammation. B2R is activated by kinins, while B1R is activated by kinins that lack the C-terminal arginine residue. Perturbations of the kinin system have been implicated in inflammation, chronic pain, vasculopathy, neuropathy, obesity, diabetes, and cancer. In general, excess activation and signaling of the kinin system lead to a pro-inflammatory state. Depending on the disease context, agonism or antagonism of the bradykinin receptors have been considered as therapeutic options. In this review, we summarize molecular imaging agents targeting these G protein-coupled receptors, including optical and radioactive probes that have been used to interrogate B1R/B2R expression at the cellular and anatomical levels, respectively. Several of these preclinical agents, described herein, have the potential to guide therapeutic interventions for these receptors.

Evaluation of Met-Val-Lys as a Renal Brush Border Enzyme-Cleavable Linker to Reduce Kidney Uptake of 68Ga-Labeled DOTA-Conjugated Peptides and Peptidomimetics.

High kidney uptake is a common feature of peptide-based radiopharmaceuticals, leading to reduced detection sensitivity for lesions adjacent to kidneys and lower maximum tolerated therapeutic dose. In this study, we evaluated if the Met-Val-Lys (MVK) linker could be used to lower kidney uptake of 68Ga-labeled DOTA-conjugated peptides and peptidomimetics. A model compound, [68Ga]Ga-DOTA-AmBz-MVK(Ac)-OH (AmBz: aminomethylbenzoyl), and its derivative, [68Ga]Ga-DOTA-AmBz-MVK(HTK01166)-OH, coupled with the PSMA (prostate-specific membrane antigen)-targeting motif of the previously reported HTK01166 were synthesized and evaluated to determine if they could be recognized and cleaved by the renal brush border enzymes. Additionally, positron emission tomography (PET) imaging, ex vivo biodistribution and in vivo stability studies were conducted in mice to evaluate their pharmacokinetics. [68Ga]Ga-DOTA-AmBz-MVK(Ac)-OH was effectively cleaved specifically by neutral endopeptidase (NEP) of renal brush border enzymes at the Met-Val amide bond, and the radio-metabolite [68Ga]Ga-DOTA-AmBz-Met-OH was rapidly excreted via the renal pathway with minimal kidney retention. [68Ga]Ga-DOTA-AmBz-MVK(HTK01166)-OH retained its PSMA-targeting capability and was also cleaved by NEP, although less effectively when compared to [68Ga]Ga-DOTA-AmBz-MVK(Ac)-OH. The kidney uptake of [68Ga]Ga-DOTA-AmBz-MVK(HTK01166)-OH was 30% less compared to that of [68Ga]Ga-HTK01166. Our data demonstrated that derivatives of [68Ga]Ga-DOTA-AmBz-MVK-OH can be cleaved specifically by NEP, and therefore, MVK can be a promising cleavable linker for use to reduce kidney uptake of radiolabeled DOTA-conjugated peptides and peptidomimetics.

Synthesis and evaluation of bifunctional tetrahydroxamate chelators for labeling antibodies with 89Zr for imaging with positron emission tomography.

Two novel bifunctional tetrahydroxamate chelators 3 and 4 were synthesized and evaluated for labeling antibodies with 89Zr for positron emission tomography imaging. Compared to previously reported tetrahydroxamate chelators 1 and 2 with an iminodiacetamide backbone, 3 and 4 were based on an extended iminodipropionamide and dipropylenetriamine backbone, respectively. Trastuzumab conjugates of 3 and 4 were efficiently labeled with 89Zr (>95% radiochemical yield). The in vitro plasma stability of 89Zr-4-Trastuzumab and especially 89Zr-3-Trastuzumab was greatly improved over previously reported 89Zr-1-Trastuzumab and 89Zr-2-Trastuzumab, but their demetalation remained higher and faster than 89Zr-deferoxamine (DFO)-Trastuzumab. These observations were confirmed by PET imaging and biodistribution in mice, with significant higher bone uptake for 89Zr-4-Trastuzumab, followed by 89Zr-3-Trastuzumab, and to a lesser extent for 89Zr-DFO-Trastuzumab. Molecular modeling showed that 3 and 4 with an extended backbone could form eight-coordinate Zr-complexes as compared to only seven-coordinate Zr-complexes of 1 and 2. Our data suggest further elongation of linker length between hydroxamate motifs of this class of chelators is needed to reach a better Zr-coordination configuration and improve in vivo stability.

89Zr for antibody labeling and in vivo studies - A comparison between liquid and solid target production.

INTRODUCTION: Zirconium-89 (89Zr, t1/2=78.4h) liquid target (LT) production offers an approach to introduce this positron-emitting isotope to cyclotron centres without the need for a separate solid target (ST) production set up. We compared the production, purification, and antibody radiolabeling yields of 89Zr-(LT) and 89Zr-(ST), and assessed the feasibility of 89Zr-(LT) for preclinical PET/CT. METHODS: 89Zr-(ST) production was performed with an 89Y foil on a TR 19 cyclotron at 13.8MeV. For LT production; an aqueous solution of yttrium nitrate (Y(NO3)3·6H2O) was irradiated on a TR 13 cyclotron at 12MeV. 89Zr was purified from the ST or LT material with hydroxamate resin, and used to radiolabel p-SCN-Bn-Deferoxamine (DFO)-conjugated Trastuzumab. MicroPET-CT imaging was performed at 1, 3 and 5days post-injection of 89Zr-DFO-Trastuzumab from ST or LT with biodistribution analysis on day 5. RESULTS: Irradiation of the ST yielded 2.88±1.07GBq/µA with a beam current of 14.0±3.8µA and irradiation time of 137±48min at end of bombardment while LT yielded 0.27±0.05GBq/µA with a beam current of 9.9±2.2µA and irradiation time of 221±29min. Radiolabeling of DFO-Trastuzumab with 89Zr-(ST) or 89Zr-(LT) was successful with purity>97% and specific activity>0.12MBq/µg (of antibody). MicroPET-CT imaging and biodistribution profiles showed similar uptake of 89Zr-(ST)-DFO-Trastuzumab and 89Zr-(LT)-DFO-Trastuzumab in tumor and all organs of interest. CONCLUSION: 89Zr-(LT) was effectively used to prepare antibody bioconjugates with specific activities suitable for small animal imaging. PET imaging and biodistribution revealed similar behaviours between bioconjugates labeled with 89Zr produced from the two target systems. ADVANCES IN KNOWLEDGE AND IMPLICATIONS FOR PATIENT CARE: These results have important implications for the production of PET isotopes such as 89Zr to cyclotron facilities with only LT capabilities - such as most clinical centres - expanding the availability of 89Zr-immunoPET.

Synthesis and evaluation of an 18 F-labeled trifluoroborate derivative of 2-nitroimidazole for imaging tumor hypoxia with positron emission tomography.

2-Nitroimidazole-based hypoxia imaging tracers such as 18 F-FMISO are normally imaged at late time points (several hours post-injection) due to their slow clearance from background tissues. Here, we investigated if a hydrophilic zwitterion-based ammoniomethyl-trifluoroborate derivative of 2-nitroimidazole, 18 F-AmBF3 -Bu-2NI, could have the potential to image tumor hypoxia at earlier time points. AmBF3 -Bu-2NI was prepared in 4 steps. 18 F labeling was conducted via 18 F-19 F isotope exchange reaction, and 18 F-AmBF3 -Bu-2NI was obtained in 14.8 ± 0.4% (n = 3) decay-corrected radiochemical yield with 24.5 ± 5.2 GBq/µmol specific activity and >99% radiochemical purity. Imaging and biodistribution studies in HT-29 tumor-bearing mice showed that 18 F-AmBF3 -Bu-2NI cleared quickly from blood and was excreted via the hepatobiliary and renal pathways. However, the tumor was not visualized in PET images until 3 hours post-injection due to low tumor uptake (0.54 ± 0.13 and 0.19 ± 0.04%ID/g at 1 and 3 hours post-injection, respectively). The low tumor uptake is likely due to the highly hydrophilic motif of ammoniomethyl-trifluoroborate that prevents free diffusion of 18 F-AmBF3 -Bu-2NI across the cell membrane. Our results suggest that highly hydrophilic 18 F-labeled ammoniomethyl-trifluoroborate derivatives might not be suitable for imaging intracellular targets including nitroreductase, a common tumor hypoxia imaging target.

p-NO2-Bn-H4neunpa and H4neunpa-Trastuzumab: Bifunctional Chelator for Radiometalpharmaceuticals and 111In Immuno-Single Photon Emission Computed Tomography Imaging.

Potentially nonadentate (N5O4) bifunctional chelator p-SCN-Bn-H4neunpa and its immunoconjugate H4neunpa-trastuzumab for 111In radiolabeling are synthesized. The ability of p-SCN-Bn-H4neunpa and H4neunpa-trastuzumab to quantitatively radiolabel 111InCl3 at an ambient temperature within 15 or 30 min, respectively, is presented. Thermodynamic stability determination with In3+, Bi3+, and La3+ resulted in high conditional stability constant (pM) values. In vitro human serum stability assays have demonstrated both 111In complexes to have high stability over 5 days. Mouse biodistribution of [111In][In(p-NO2-Bn-neunpa)]-, compared to that of [111In][In(p-NH2-Bn-CHX-A″-diethylenetriamine pentaacetic acid (DTPA))]2-, at 1, 4, and 24 h shows fast clearance of both complexes from the mice within 24 h. In a second mouse biodistribution study, the immunoconjugates 111In-neunpa-trastuzumab and 111In-CHX-A″-DTPA-trastuzumab demonstrate a similar distribution profile but with slightly lower tumor uptake of 111In-neunpa-trastuzumab compared to that of 111In-CHX-A″-DTPA-trastuzumab. These results were also confirmed by immuno-single photon emission computed tomography (immuno-SPECT) imaging in vivo. These initial investigations reveal the acyclic bifunctional chelator p-SCN-Bn-H4neunpa to be a promising chelator for 111In (and other radiometals) with high in vitro stability and also show H4neunpa-trastuzumab to be an excellent 111In chelator with promising biodistribution in mice.

Evaluation of agonist and antagonist radioligands for somatostatin receptor imaging of breast cancer using positron emission tomography.

BACKGROUND: The somatostatin receptor subtype 2 (sstr2) is expressed on a majority of luminal breast cancers, however SPECT and scintigraphy imaging with agonistic sstr2 probes has been sub-optimal. High affinity antagonists can access more binding sites on the cell surface, resulting in higher tumor uptake and improved sensitivity. We compared the tumor uptake and biodistribution of the antagonist 68Ga-NODAGA-JR11 with two agonists 68Ga-DOTA-Tyr3-octreotide (68Ga-DOTATOC) and 68Ga-DOTA-Tyr3-octreotate (68Ga-DOTATATE), in the human, sstr2-positive, luminal breast cancer model: ZR-75-1. RESULTS: Peptides were assayed for binding affinity using a filtration-based competitive assay to sstr2. natGa-DOTATOC and natGa-DOTATATE had excellent affinity (inhibition constant Ki: 0.9 ± 0.1 nM and 1.4 ± 0.3 nM respectively) compared to natGa-NODAGA-JR11 (25.9 ± 0.2 nM). The number of binding sites on ZR-75-1 cells was determined in vitro by saturation assays. Agonist 67/natGa-DOTATOC bound to 6.64 ± 0.39 × 104 sites/cells, which was 1.5-fold higher than 67/natGa-NODAGA-JR11 and 2.3-fold higher than 67/natGa-DOTATATE. All three 68Ga-labeled peptides were obtained in good decay-corrected radiochemical yield (61-68%) and were purified by high performance liquid chromatography to ensure high specific activity (137 - 281 MBq/nmol at the end of synthesis). NOD scid gamma mice bearing ZR-75-1 tumors were injected intravenously with the labeled peptides and used for PET/CT imaging and biodistribution at 1 h post-injection. We found that 68Ga-DOTATOC had the highest tumor uptake (18.4 ± 2.9%ID/g), followed by 68Ga-DOTATATE (15.2 ± 2.2%ID/g) and 68Ga-NODAGA-JR11 (12.2 ± 0.8%ID/g). Tumor-to-blood and tumor-to-muscle ratios were also higher for the agonists (>40 and >150 respectively), compared to the antagonist (15.6 ± 2.2 and 45.2 ± 11.6 respectively). CONCLUSIONS: The antagonist 68Ga-NODAGA-JR11 had the lowest tumor uptake and contrast compared to agonists 68Ga-DOTATOC and 68Ga-DOTATATE in ZR-75-1 xenografts.The main contributing factor to this result could be the use of an endogenously expressing cell line, which may differ from previously published transfected models in the number of low-affinity, antagonist-specific binding sites. The relative merit of agonists versus antagonists for sstr2 breast cancer imaging warrants further investigation, first in preclinical models with other sstr2-positive breast cancer xenografts, and ultimately in luminal breast cancer patients.

Feasibility of an emotional health curriculum for elementary school students in an underserved Hispanic community.

PROBLEM: Hispanic children have greater mental health challenges but fewer received mental health services than other ethnic groups. A classroom-based Emotional Health Curriculum (EHC) was developed to address mental health disparities in an underserved Hispanic community. METHODS: A quasi-experimental design with one group pre- and post-intervention was used to test the feasibility of an 8-week EHC for one hundred 3rd and 4th grade children in a dual-immersion Spanish-English elementary school. Limited efficacy was measured by changes in depression and anxiety scores reported by children and teachers. Acceptance was evaluated by a child-reported satisfaction survey and a focus group in which the four teachers shared their experiences. Implementation was measured by participation, retention, and fidelity rates. FINDINGS: The child-reported depression and anxiety and teacher-reported depression were significantly decreased in at-risk children with the effect size ranging from 0.60 to 1.16 (ps < 0.05). The majority of children (89.7%) enjoyed the EHC and teachers observed that children had acquired skills to manage their emotional distress. The participation, retention, and fidelity rates were 98%, 94%, and 99.13%, respectively. CONCLUSIONS: The results provide promising evidence that the EHC has the potential to improve depression and anxiety symptoms in at-risk children.

Polymorphisms of asparaginase pathway and asparaginase-related complications in children with acute lymphoblastic leukemia.

PURPOSE: Asparaginase (ASNase) is a standard and critical component in the therapy of childhood acute lymphoblastic leukemia (ALL), but it is also associated with several toxicities. EXPERIMENTAL DESIGN: We recently reported the results of an association study between ASNase pathway genes and event-free survival (EFS) in childhood patients with ALL. The same polymorphisms were interrogated here in relation to allergies, pancreatitis, and thrombotic events following treatment with E. coli ASNase. RESULTS: Among patients of the discovery group, allergies, and pancreatitis were more frequent in individuals who are homozygous for the triple-repeat allele (3R) of the asparagine synthetase (ASNS) gene, resulting in remarkably higher risk of these toxicities associated with 3R3R genotype [OR for allergies, 14.6; 95% confidence interval (CI), 3.6-58.7; P < 0.0005 and OR for pancreatitis, 8.6; 95% CI, 2.0-37.3; P = 0.01]. In contrast, the ASNS haplotype *1 harboring double-repeat (2R) allele had protective effect against these adverse reactions (P ≤ 0.01). The same haplotype was previously reported to confer reduction in EFS. The risk effect of 3R3R genotype was not replicated in the validation cohort, whereas the protective effect of haplotype *1 against allergies was maintained (P ≤ 0.002). Analysis with additional polymorphisms in ASNS locus in lymphoblastoid cell lines showed that haplotype *1 is diversified in several subtypes of which one was associated with reduced in vitro sensitivity to ASNase (rs10486009, P = 0.01) possibly explaining an association seen in clinical setting. CONCLUSIONS: This finding might have implication for treatment individualization in ALL and other cancers using asparagine depletion strategies. Clin Cancer Res; 21(2); 329-34. ©2014 AACR. See related commentary by Avramis, p. 230.

Polymorphisms of the vincristine pathway and response to treatment in children with childhood acute lymphoblastic leukemia.

BACKGROUND: Vincristine (VCR) is a standard component in the treatment of childhood acute lymphoblastic leukemia (ALL). VCR cytotoxicity is primarily due to its ability to disrupt the formation of microtubules of the mitotic spindle. PATIENTS & METHODS: Seventeen polymorphisms in regulatory and coding regions of genes controlling VCR targets (TUBB1, MAP4, ACTG1 and CAPG) or potentially influencing VCR levels (ABCB1 and CYP3A5) were investigated for an association with peripheral neuropathy and outcome in childhood ALL patients. RESULTS: High-grade neurotoxicity was more frequent in carriers of the A allele of synonymous (Ala310) G to A (rs1135989) variation in the ACTG1 gene. Substitution (rs4728709) in the promoter of the ABCB1 gene had a protective effect against lower grade neurotoxicity and C to A variation (rs3770102) located 17 nucleotides upstream from the transcription start site had a protective effect against high-grade neurotoxicity. Patients with the ABCB1 3435TT genotype had lower event-free survival; the association with event-free survival was not supported by the analysis in the replication patient set. CONCLUSION: The polymorphisms in the ACTG1, CAPG and ABCB1 genes may modulate VCR-related neurotoxicity, whereas the risk of relapse seems not to be affected by the genes of the VCR pathway.

A compartmental model of mouse thrombopoiesis and erythropoiesis to predict bone marrow toxicity after internal irradiation.

UNLABELLED: In targeted radionuclide radiotherapy, the relationship between bone marrow (BM) toxicity and absorbed dose seems to be elusive. A compartmental model of mouse thrombopoiesis and erythropoiesis was set up to predict the depletion of hematopoietic cells as a function of the irradiation dose delivered to BM by injected radiopharmaceuticals. All simulated kinetics were compared with experimental toxicity for several stages of differentiation of the 2 hematopoietic lineages. METHODS: C57BL/6 mice were injected either with (18)FNa (37 and 60 MBq), a bone-seeking agent, or with saline. BM mean absorbed doses were calculated according to the MIRD formalism from small-animal PET/CT images. Hematologic toxicity was monitored over time, after (18)FNa injection, by studying BM progenitors and precursors in addition to blood cells. The compartmental model takes into account the pharmacokinetics of the compound, in addition to cellular kinetics and cell radiosensitivities for the 2 studied lineages. RESULTS: Because biodistribution studies showed an uptake of (18)FNa in bones, the skeleton was considered as the principal source organ of BM irradiation. The time-activity curve obtained from validated quantification of PET/CT images allowed for the calculation of mean absorbed doses to the whole BM of 2.1 and 3.4 Gy for (18)FNa injections of 37 and 60 MBq, respectively. Concerning hematologic toxicity, the model was in good agreement for the 2 absorbed doses with experimental measurements of cell depletion for platelets, progenitors, and precursors within the BM in terms of time to nadir, depletion intensity, and time to recovery. The same agreement was obtained for red blood cells and their precursors. Model predictions demonstrated that BM toxicity was in correlation with the mean absorbed dose as higher depletions at nadir and longer delays to recovery were noticed for 3.4 Gy than for 2.1 Gy. CONCLUSION: The developed compartmental model of thrombopoiesis and erythropoiesis in a BM toxicity context, after internal irradiation, allowed for the prediction of cell kinetics of BM progenitors, precursors, and mature blood cells in a dose-dependent manner. This model could therefore be used to predict hematologic toxicity in preclinical internal radiotherapy to study the dose-response relationship.

Allele-specific Col1a1 silencing reduces mutant collagen in fibroblasts from Brtl mouse, a model for classical osteogenesis imperfecta.

Gene silencing approaches have the potential to become a powerful curative tool for a variety of monogenic diseases caused by gain-of-function mutations. Classical osteogenesis imperfecta (OI), a dominantly inherited bone dysplasia, is characterized in its more severe forms by synthesis of structurally abnormal type I collagen, which exerts a negative effect on extracellular matrix. Specific suppression of the mutant (Mut) allele would convert severe OI forms to the mild type caused by a quantitative defect in normal collagen. Here, we describe the in vitro and ex vivo investigation of a small interfering RNA (siRNA) approach to allele-specific gene silencing using Mut Col1a1 from the Brtl mouse, a well-characterized model for classical human OI. A human embryonic kidney cell line, which expresses the firefly luciferase gene, combined with either wild-type or Mut Brtl Col1a1 exon 23 sequences, was used for the first screening. The siRNAs selected based on their specificity and the corresponding short hairpin RNAs (shRNAs) subcloned in a lentiviral vector were evaluated ex vivo in Brtl fibroblasts for their effect on collagen transcripts and protein. A preferential reduction of the Mut allele of up to 52% was associated with about 40% decrease of the Mut protein, with no alteration of cell proliferation. Interestingly, a downregulation of HSP47, a specific collagen chaperone known to be upregulated in some OI cases, was detected. Our data support further testing of shRNAs and their delivery by lentivirus as a strategy to specifically suppress the Mut allele in mesenchymal stem cells of OI patients for autologous transplantation.

Bim polymorphisms: influence on function and response to treatment in children with acute lymphoblastic leukemia.

PURPOSE: Corticosteroids induce apoptosis in the malignant lymphoid cells and are critical component of combination therapy for acute lymphoblastic leukemia (ALL). Several genome-wide microarray studies showed major implication of proapoptotic Bim in mediating corticosteroid-related resistance in leukemia cells. EXPERIMENTAL DESIGN: We investigated Bim gene polymorphisms and their association with childhood ALL outcome, and the mechanism underlying the observed finding. RESULTS: Lower overall survival (OS) was associated with Bim C29201T located in Bcl-2 homology 3 (BH3) domain (P = 0.01). An association remained significant in multivariate model (P = 0.007), was more apparent in high-risk patients (P = 0.004) and patients treated with dexamethasone (P = 0.009), and was subsequently confirmed in the replication patient cohort (P = 0.03). RNA analysis revealed that C29201T affects generation of γ isoforms (γ1) that lack proapoptotic BH3 domain. The phenotypic effect was minor suggesting the influence of additional factors that may act in conjunction with Bim genotype. Combined analysis with Mcl gene polymorphism (G-486T) revealed profound reduction in OS in individuals with both risk genotypes (P < 0.0005 in discovery and P = 0.002 in replication cohort) and particularly in high-risk patients (P ≤ 0.008). CONCLUSIONS: Increased expression of prosurvival Mcl1 and presence of Bim isoforms lacking proapoptotic function might explain marked reduction of OS in a disease and dose-dependent manner in ALL patients carrying Bim- and Mcl1-risk genotypes.