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Improvements in the diagnosis and treatment of systemic breast cancer have led to a prolongation in patient survival. Unfortunately, these advances are also associated with an increased incidence of brain metastases (BM), with the result that many patients succumb due to BM treatment failure. Intracranial delivery of many chemotherapeutic agents and other therapeutics is hindered by the presence of an impermeable blood-brain barrier (BBB) designed to protect the brain from harmful substances. The formation of BM compromises the integrity of the BBB, resulting in a highly heterogeneous blood-tumor barrier (BTB) with varying degrees of vascular permeability. Here, we discuss how blood vessels play an important role in the formation of brain micrometastases as well as in the transformation from poorly permeable BM to highly permeable BM. We then review the role of BTB vascular permeability in the diagnostics and the choice of treatment regimens for breast cancer brain metastases (BCBM) and discuss whether the vasculature of primary breast cancers can serve as a biomarker for BM. Specifically, we examine the association between the vascular permeability of BCBM and their accumulation of large molecules such as antibodies, which remains largely unexplored.

The promising characteristics of the 7.2-h radiohalogen (211)At have long been recognized; including having chemical properties suitable for labeling targeting vectors ranging from small organic molecules to proteins, and the emission of only one α-particle per decay, providing greater control over off-target effects. Unfortunately, the impact of (211)At within the targeted α-particle therapy domain has been constrained by its limited availability. Paradoxically, the most commonly used production method – via the (209)Bi(α,2n)(211)At reaction – utilizes a widely available natural material (bismuth) as the target and straightforward cyclotron irradiation methodology. On the other hand, the most significant impediment to widespread (211)At availability is the need for an accelerator capable of generating ≥28 MeV α-particles with sufficient beam intensities to make clinically relevant levels of (211)At. In this review, current methodologies for the production and purification of (211)At – both by the direct production route noted above and via a (211)Rn generator system – will be discussed. The capabilities of cyclotrons that currently produce (211)At will be summarized and the characteristics of other accelerators that could be utilized for this purpose will be described. Finally, the logistics of networks, both academic and commercial, for facilitating (211)At distribution to locations remote from production sites will be addressed.

Purpose: Camelid single-domain antibody-fragments (sdAb) have beneficial pharmacokinetic properties, and those targeted to HER2 can be used for imaging of HER2-overexpressing cancer. Labeled with a therapeutic radionuclide, they may be used for HER2-targeted therapy. Here, we describe the generation of a 131I-labeled sdAb as a theranostic drug to treat HER2-overexpressing cancer.Experimental Design: Anti-HER2 sdAb 2Rs15d was labeled with 131I using [131I]SGMIB and evaluated in vitro. Biodistribution was evaluated in two HER2+ murine xenograft models by micro-SPECT/CT imaging and at necropsy, and under challenge with trastuzumab and pertuzumab. The therapeutic potential of [131I]SGMIB-2Rs15d was investigated in two HER2+ tumor mouse models. A single-dose toxicity study was performed in mice using unlabeled [127I]SGMIB-sdAb at 1.4 mg/kg. The structure of the 2Rs15d–HER2 complex was determined by X-ray crystallography.Results: [131I]SGMIB-2Rs15d bound specifically to HER2+ cells (Kd = 4.74 ± 0.39 nmol/L). High and specific tumor uptake was observed in both BT474/M1 and SKOV-3 tumor xenografted mice and surpassed kidney levels by 3 hours. Extremely low uptake values were observed in other normal tissues at all time points. The crystal structure revealed that 2Rs15d recognizes HER2 Domain 1, consistent with the lack of competition with trastuzumab and pertuzumab observed in vivo. [131I]SGMIB-2Rs15d alone, or in combination with trastuzumab, extended median survival significantly. No toxicity was observed after injecting [127I]SGMIB-2Rs15d.Conclusions: These findings demonstrate the theranostic potential of [131I]SGMIB-2Rs15d. An initial scan using low radioactive [*I]SGMIB-2Rs15d allows patient selection and dosimetry calculations for subsequent therapeutic [131I]SGMIB-2Rs15d and could thereby impact therapy outcome on HER2+ breast cancer patients. Clin Cancer Res; 23(21); 6616–28. ©2017 AACR.

D2C7-(scdsFv)-PE38KDEL (D2C7-IT) is a novel immunotoxin that reacts with wild-type epidermal growth factor receptor (EGFRwt) and mutant EGFRvIII proteins overexpressed in glioblastomas. This study assessed the toxicity of intracerebral administration of D2C7-IT to support an initial Food and Drug Administration Investigational New Drug application. After the optimization of the formulation and administration, two cohorts (an acute and chronic cohort necropsied on study days 5 and 34) of Sprague-Dawley (SD) rats (four groups of 5 males and 5 females) were infused with the D2C7-IT formulation at total doses of 0, 0.05, 0.1, 0.4 µg (the acute cohort) and 0, 0.05, 0.1, 0.35 µg (the chronic cohort) for approximately 72 hours by intracerebral convection-enhanced delivery using osmotic pumps. Mortality was observed in the 0.40 µg (5/10 rats) and 0.35 µg (4/10 rats) high-dose groups of each cohort. Body weight loss and abnormal behavior were only revealed in the rats treated with high doses of D2C7-IT. No dose-related effects were observed in clinical laboratory tests in either cohort. A gross pathologic examination of systemic tissues from the high-dose and control groups in both cohorts exhibited no dose-related or drug-related pathologic findings. Brain histopathology revealed the frequent occurrence of dose-related encephalomalacia, edema, and demyelination in the high-dose groups of both cohorts. In this study, the maximum tolerated dose of D2C7-IT was determined to be between 0.10 and 0.35 µg, and the no observed adverse effect level was 0.05 µg in SD rats. Both parameters were utilized to design the Phase I/II D2C7-IT clinical trial.

INTRODUCTION: Radioiodinated meta-iodobenzylguanidine (MIBG), a norepinephrine transporter (NET) substrate, has been extensively used as an imaging agent to study the pathophysiology of the heart and for the diagnosis and treatment of neuroendocrine tumors. The goal of this study was to develop an (18)F-labeled analogue of MIBG that like MIBG itself could be synthesized in a single radiochemical step. Towards this end, we designed 4-fluoropropoxy-3-iodobenzylguanidine (FPOIBG). METHODS: Standards of FPOIBG and 4-fluoropropoxy-3-bromobenzylguanidine (FPOBBG) as well as their tosylate precursors for labeling with (18)F, and a tin precursor for the preparation of radioiodinated FPOIBG were synthesized. Radiolabeled derivatives were synthesized by nucleophilic substitution and electrophilic iododestannylation from the corresponding precursors. Labeled compounds were evaluated for NET transporter recognition in in vitro assays using three NET-expressing cell lines and in biodistribution experiments in normal mice, with all studies performed in a paired-label format. Competitive inhibition of [(125)I]MIBG uptake by unlabeled benzylguanidine compounds was performed in UVW-NAT cell line to determine IC50 values. RESULTS: [(18)F]FPOIBG was synthesized from the corresponding tosylate precursor in 5.2 ± 0.5% (n = 6) overall radiochemical yields starting with aqueous fluoride in about 105 min. In a paired-label in vitro assay, the uptake of [(18)F]FPOIBG at 2h was 10.2 ± 1.5%, 39.6 ± 13.4%, and 13.3 ± 2.5%, in NET-expressing SK-N-SH, UVW-NAT, and SK-N-BE(2c) cells, respectively, while these values for [(125)I]MIBG were 57.3 ± 8.1%, 82.7 ± 8.9%, and 66.3 ± 3.6%. The specificity of uptake of both tracers was demonstrated by blocking with desipramine. The (125)I-labeled congener of FPOIBG gave similar results. On the other hand, [(18)F]FPOBBG, a compound recently reported in the literature, demonstrated much higher uptake, albeit less than that of co-incubated [(125)I]MIBG. IC50 values for FPOIBG were higher than those obtained for MIBG and FPOBBG. Unlike the case with [(18)F]FPOBBG, the heart uptake [(18)F]FPOIBG in normal mice was significantly lower than that of MIBG. CONCLUSION: Although [(18)F]FPOIBG does not appear to warrant further consideration as an (18)F-labeled MIBG analogue, analogues wherein the iodine in it is replaced with a chlorine, fluorine or hydrogen might be worth pursuing. ADVANCES IN KNOWLEDGE AND IMPLICATIONS FOR PATIENT CARE: An (18)F-labeled analogue of the well-known radiopharmaceutical MIBG could have significant impact, potentially improving imaging of NET related disease in cardiology and in the imaging of neuroendocrine tumors. Although (18)F-labeled analogues of MIBG have been reported including LMI1195, we undertook this work hypothesizing that based on its greater structural similarity to MIBG, FPOIBG might be a better analogue than LMI1195.

PURPOSE: Our previous studies with F-18-labeled anti-HER2 single-domain antibodies (sdAbs) utilized 5F7, which binds to the same epitope on HER2 as trastuzumab, complicating its use for positron emission tomography (PET) imaging of patients undergoing trastuzumab therapy. On the other hand, sdAb 2Rs15d binds to a different epitope on HER2 and thus might be a preferable vector for imaging in these patients. The aim of this study was to evaluate the tumor targeting of F-18 -labeled 2Rs15d in HER2-expressing breast carcinoma cells and xenografts. PROCEDURES: sdAb 2Rs15d was labeled with the residualizing labels N-succinimidyl 3-((4-(4-[(18)F]fluorobutyl)-1H-1,2,3-triazol-1-yl)methyl)-5-(guanidinomethyl)benzoate ([(18)F]RL-I) and N-succinimidyl 4-guanidinomethyl-3-[(125)I]iodobenzoate ([(125)I]SGMIB), and the purity and HER2-specific binding affinity and immunoreactivity were assessed after labeling. The biodistribution of I-125- and F-18-labeled 2Rs15d was determined in SCID mice bearing subcutaneous BT474M1 xenografts. MicroPET/x-ray computed tomograph (CT) imaging of [(18)F]RL-I-2Rs15d was performed in this model and compared to that of nonspecific sdAb [(18)F]RL-I-R3B23. MicroPET/CT imaging was also done in an intracranial HER2-positive breast cancer brain metastasis model after administration of 2Rs15d-, 5F7-, and R3B23-[(18)F]RL-I conjugates. RESULTS: [(18)F]RL-I was conjugated to 2Rs15d in 40.8 ± 9.1 % yield and with a radiochemical purity of 97-100 %. Its immunoreactive fraction (IRF) and affinity for HER2-specific binding were 79.2 ± 5.4 % and 7.1 ± 0.4 nM, respectively. [(125)I]SGMIB was conjugated to 2Rs15d in 58.4 ± 8.2 % yield and with a radiochemical purity of 95-99 %; its IRF and affinity for HER2-specific binding were 79.0 ± 12.9 % and 4.5 ± 0.8 nM, respectively. Internalized radioactivity in BT474M1 cells in vitro for [(18)F]RL-I-2Rs15d was 43.7 ± 3.6, 36.5 ± 2.6, and 21.7 ± 1.2 % of initially bound radioactivity at 1, 2, and 4 h, respectively, and was similar to that seen for [(125)I]SGMIB-2Rs15d.Uptake of [(18)F]RL-I-2Rs15d in subcutaneous xenografts was 16-20 %ID/g over 1-3 h. Subcutaneous tumor could be clearly delineated by microPET/CT imaging with [(18)F]RL-I-2Rs15d but not with [(18)F]RL-I-R3B23. Intracranial breast cancer brain metastases could be visualized after intravenous administration of both [(18)F]RL-I-2Rs15d and [(18)F]RL-I-5F7. CONCLUSIONS: Although radiolabeled 2Rs15d conjugates exhibited lower tumor cell retention both in vitro and in vivo than that observed previously for 5F7, given that it binds to a different epitope on HER2 from those targeted by the clinically utilized HER2-targeted therapeutic antibodies trastuzumab and pertuzumab, F-18-labeled 2Rs15d has potential for assessing HER2 status by PET imaging after trastuzumab and/or pertuzumab therapy.

Intratumoral radiation therapy – ‘brachytherapy’ – is a highly effective treatment for solid tumors, particularly prostate cancer. Current titanium seed implants, however, are permanent and are limited in clinical application to indolent malignancies of low- to intermediate-risk. Attempts to develop polymeric alternatives, however, have been plagued by poor retention and off-target toxicity due to degradation. Herein, we report on a new approach whereby thermally sensitive micelles composed of an elastin-like polypeptide (ELP) are labeled with the radionuclide 131I to form an in situ hydrogel that is stabilized by two independent mechanisms: first, body heat triggers the radioactive ELP micelles to rapidly phase transition into an insoluble, viscous coacervate in under 2 minutes; second, the high energy β-emissions of 131I further stabilize the depot by introducing crosslinks within the ELP depot over 24 hours. These injectable brachytherapy hydrogels were used to treat two aggressive orthotopic tumor models in athymic nude mice: a human PC-3M-luc-C6 prostate tumor and a human BxPc3-luc2 pancreatic tumor model. The ELP depots retained greater than 52% and 70% of their radioactivity through 60 days in the prostate and pancreatic tumors with no appreciable radioactive accumulation (≤ 0.1% ID) in off-target tissues after 72 hours. The 131I-ELP depots achieved >95% tumor regression in the prostate tumors (n=8); with a median survival of more than 60 days compared to 12 days for control mice. For the pancreatic tumors, ELP brachytherapy (n=6) induced significant growth inhibition (p = 0.001, ANOVA) and enhanced median survival to 27 days over controls.

Alpha-particle emitters have a high linear energy transfer and short range, offering the potential for treating micrometastases while sparing normal tissues. We developed a urea-based, 211At-labeled small molecule targeting prostate-specific membrane antigen (PSMA) for the treatment of micrometastases due to prostate cancer (PC). Methods: PSMA-targeted (2S)-2-(3-(1-carboxy-5-(4-211At-astatobenzamido)pentyl)ureido)-pentanedioic acid (211At-6) was synthesized. Cellular uptake and clonogenic survival were tested in PSMA-positive (PSMA+) PC3 PIP and PSMA-negative (PSMA−) PC3 flu human PC cells after 211At-6 treatment. The antitumor efficacy of 211At-6 was evaluated in mice bearing PSMA+ PC3 PIP and PSMA– PC3 flu flank xenografts at a 740-kBq dose and in mice bearing PSMA+, luciferase-expressing PC3-ML micrometastases. Biodistribution was determined in mice bearing PSMA+ PC3 PIP and PSMA– PC3 flu flank xenografts. Suborgan distribution was evaluated using α-camera images, and microscale dosimetry was modeled. Long-term toxicity was assessed in mice for 12 mo. Results: 211At-6 treatment resulted in PSMA-specific cellular uptake and decreased clonogenic survival in PSMA+ PC3 PIP cells and caused significant tumor growth delay in PSMA+ PC3 PIP flank tumors. Significantly improved survival was achieved in the newly developed PSMA+ micrometastatic PC model. Biodistribution showed uptake of 211At-6 in PSMA+ PC3 PIP tumors and in kidneys. Microscale kidney dosimetry based on α-camera images and a nephron model revealed hot spots in the proximal renal tubules. Long-term toxicity studies confirmed that the dose-limiting toxicity was late radiation nephropathy. Conclusion: PSMA-targeted 211At-6 α-particle radiotherapy yielded significantly improved survival in mice bearing PC micrometastases after systemic administration. 211At-6 also showed uptake in renal proximal tubules resulting in late nephrotoxicity, highlighting the importance of long-term toxicity studies and microscale dosimetry.

Convection-enhanced delivery (CED) permits site-specific therapeutic drug delivery within interstitial spaces at increased dosages through circumvention of the blood-brain barrier. CED is currently limited by suboptimal methodologies for monitoring the delivery of therapeutic agents that would permit technical optimization and enhanced therapeutic efficacy.To determine whether a readily available small-molecule MRI contrast agent, gadolinium-diethylene triamine pentaacetic acid (Gd-DTPA), could effectively track the distribution of larger therapeutic agents.Gd-DTPA was coinfused with the larger molecular tracer, I-labeled human serum albumin (I-HSA), during CED of an EGFRvIII-specific immunotoxin as part of treatment for a patient with glioblastoma.Infusion of both tracers was safe in this patient. Analysis of both Gd-DTPA and I-HSA during and after infusion revealed a high degree of anatomical and volumetric overlap.Gd-DTPA may be able to accurately demonstrate the anatomic and volumetric distribution of large molecules used for antitumor therapy with high resolution and in combination with fluid-attenuated inversion recovery (FLAIR) imaging, and provide additional information about leaks into cerebrospinal fluid spaces and resection cavities. Similar studies should be performed in additional patients to validate our findings and help refine the methodologies we used.

Pancreatic tumors remain one of the deadliest manifestations of cancer in clinical oncology. Even when tumors are diagnosed early, remain loco-regionalized, and receive cutting-edge cancer treatment, the 5-year survival rate remains less than 27%. This high mortality rate arises from the significant biologic barriers that inhibit drug delivery effectiveness in pancreatic tumors, as well as their underlying genetic mutations that confer resistance to traditional radiation and chemotherapeutic standards of care. In this work, we hypothesized that a new form of radiation brachytherapy might prove highly synergistic for overcoming the resistance native to pancreatic tumors. Utilizing the thermally stabilized biopolymer ELP, 131iodine radionuclides can be delivered intratumorally to provide continuous, focalized, high-dose β-radiation to treat pancreatic cancer from the inside out. Dose-escalation studies revealed that the two important variables for improving outcomes were the 131I-ELP radioactivity dose and the duration of concurrent paclitaxel therapy. 100% tumor regression was achieved across multiple genotypic models of pancreatic cancer models: MIA PaCa-2, AsPc-1, and BxPc3 (both subcutaneous and orthotopic). The potent synergistic results of 131I-ELP brachytherapy were replicated using both a novel micelle formulation of paclitaxel and the clinical standard Abraxane. Multigroup analysis revealed true molecular synergy as assessed by the Bliss Independence framework (p < 0.01). External beam irradiation, when similarly employed as comparison, merely induced growth inhibition when combined with paclitaxel utilizing a 5Gy hypofractionation regimen. Immunohistochemical analysis of treated tumors revealed differential modulation of biomolecular components that regulate the effectiveness of drug delivery and radiotoxicity. Levels of PECAM, VE-cadherin, claudin-4, HIF-1α, and stromal collagen were all specifically investigated, while macroscopic analysis revealed significantly enhanced accumulation and retention of fluorescently tagged paclitaxel when tumors were treated with biopolymer brachytherapy. Standard x-ray therapy indicated little to no improvement. No signs of any toxic side effects were observed in serologic enzyme analysis and multiorgan biodistribution studies. Citation Format: Jeffrey L. Schaal, Jayanta Bhattacharyya, Xinghai Li, Samagya Banskota, Jeremy Brownstein, David Kirsch, Michael Zalutsky, Ashutosh Chilkoti. Biopolymer β-brachytherapy delivered concomitantly with systemic paclitaxel outperforms traditional x-ray radiation to induce complete regression in multiple pancreatic tumor xenograft models [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5831.

160 Background: Radium-223 is a targeted alpha therapy that improves survival in men with mCRPC. The biologic basis for radium-223 efficacy is not completely understood. We hypothesized that PC osteomimicry, a form of epithelial plasticity leading to an osteoblastic phenotype, may contribute to the intralesional deposition of radium-223 and subsequent irradiation of the tumor microenvironment. Methods: We conducted a pharmacodynamic study of radium-223 in men with bone predominant mCRPC to investigate genomic and phenotypic alterations in circulating tumor cells (CTCs), ctDNA, and metastases. Prior to radium and 3 and 6 months after radium, liquid and metastatic biopsies were collected, including CTCs for phenotypic characterization and CTC/ctDNA genomic analysis. The primary objective was to describe the prevalence of CTC bone alkaline phosphatase (BAP) over time. We measured radium-223 decay products in tumor and surrounding normal bone during treatment. Results: We enrolled 20 men with heavily pre-treated symptomatic bone predominant mCRPC and treated with radium-223 over a median of 6 doses. 55% had elevated serum BAP. PFS was 5.5 mo; OS was 13.3 mo; 10% had unfavorable (≥5) to favorable ( < 5) Cellsearch CTC conversion; 5% had ≥30% PSA decline. We found evidence of persistent CTC BAP expression in both EpCAM+ and EpCAM- CTCs in the majority of men over time during radium treatment. We identified genomic gain of key osteomimicry regions in CTC DNA, including loci for BAP, osteopontin, and OB-cadherin. Radium-223 uptake was observed in tumor to a greater degree than surrounding normal bone. CTC DNA and matched ctDNA and CTC cultures suggested persistence of CTCs with aggressive genomic alterations such as AR, FOXA1, and MYC gain and PTEN and RB1 loss. We established multiple CTC cultures and one CTC PDX; cells exhibited evidence of epithelial plasticity with BAP expression. Conclusions: Osteomimicry may contribute to the uptake of Radium-223 within bone metastases and may thereby enhance the therapeutic benefit of radium-223. We found genomic and phenotypic evidence of osteomimicry in CTCs and CTC cultures from men with mCRPC. Clinical trial information: NCT02204943.

Medulloblastoma, the most common pediatric brain tumor, is difficult to treat because conventional therapeutic approaches result in significant toxicity to normal central nervous system tissues, compromising quality of life. Given the fact that medulloblastomas express the somatostatin subtype 2 receptor, [177Lu-DOTA0,Tyr3]octreotate ([177Lu]DOTA-TATE) could be a potentially useful targeted radiotherapeutic for the treatment of this malignancy. The current study was undertaken to evaluate this possibility in preclinical models of D341 MED human medulloblastoma by comparing the properties of [177Lu]DOTA-TATE to those of glucose-[125I-Tyr3]-octreotate ([125I]Gluc-TOCA), a radiopeptide previously shown to target this cell line. In vitro assays indicated that both labeled peptides exhibited similar cell-associated and internalized radioactivity after a 30-min incubation at 37° C; however, at the end of the 4 h incubation period, the internalized radioactivity for [177Lu]DOTA-TATE (6.22 ± 0.75%) was nearly twice that for [125I]Gluc-TOCA (3.16 ± 0.27%), with similar differences seen in total cell-associated radioactivity levels. Consistent with the results from the internalization assays, results from paired-label tissue distribution studies in athymic mice with subcutaneous D341 MED medulloblastoma xenografts showed a similar degree of tumor accumulation for [177Lu]DOTA-TATE and [125I]Gluc-TOCA at early time points but by 24 h, a more than 5-fold advantage was observed for the 177Lu-labeled peptide. Tumor-to-normal tissue ratios generally were more favorable for [177Lu]DOTA-TATE at all time points, due in part to its lower accumulation in normal tissues except kidneys. Taken together, these results suggest that [177Lu]DOTA-TATE warrants further investigation as a targeted radiotherapeutic for medulloblastoma treatment.

Adults with primary malignant glioma have an unacceptably poor outcome. Although temozolomide plus radiotherapy improves survival of newly diagnosed glioblastoma multiforme (GBM) patients,1 most patients develop tumor progression within 1 – 2 years. Outcome following recurrence is poor.2 Most tumors recur at or adjacent to the site of origin, indicating that failure to eradicate local tumor growth is a major factor contributing to poor outcome.3 For this reason, we have focused on augmenting local control to improve overall outcome by administering tumor-associated radiolabeled monoclonal antibodies (mAbs) directly into spontaneous tumor cysts, surgically created resection cavities (SCRCs), the intrathecal space, and solid tumors.4–8 Tenascin, an extracellular matrix hexabrachion glycoprotein, is expressed ubiquitously in several cancers, including high-grade gliomas, but not in normal brain.9,10 mAb 81C6, a murine isotype 2b immunoglobulin G (IgG2b) that binds to an alternatively spliced region of tenascin,9–12 reacts specifically with tenascin-expressing tumors.13 When labeled with 131I, 81C6 delays tumor growth and prolongs survival in flank and intracranial human xenograft models.14,15 An initial human experience demonstrated the specificity of 131I-labeled murine 81C6 (131I-81C6) mAb compared to 125I-labeled nonspecific IgG2b mAb but also revealed limited intratumor penetration following intravenous or intraarterial administration.16 Hence, subsequent studies incorporated administration into an SCRC, tumor cyst, or intrathecal space. Prior phase I studies established the maximum tolerated dose (MTD) of 131I-81C6 injected into the SCRC of patients with newly diagnosed and recurrent malignant brain tumors to be 120 mCi and 100 mCi, respectively.7,8 Phase II studies demonstrated that patients treated with 131I-81C6 had favorable overall survival compared to established historical controls.4,5 Based on constraints of our U.S. Food and Drug Administration (FDA)-approved phase I and II studies, patients received a “fixed” dose of 131I-81C6 in which the administered level of radioactivity was the same among groups of patients. As a result of this design, 131I dosages were not adjusted to compensate for patient-specific variables such as SCRC volume and SCRC residence time. Dosimetry of patients treated on these studies revealed a wide range of radiation absorbed doses at the SCRC margin. Moreover, outcome correlated with delivered radiation dose to the SCRC. Specifically, patients who received less than 44 Gy were more likely to develop recurrent tumor, whereas those who received significantly more than 44 Gy were more likely to develop radionecrosis. Therefore, a 44-Gy boost to the SCRC margin was considered optimal.17 We now report a pilot study using a novel, patient-specific dosing strategy of 131I-81C6 designed to achieve a 44-Gy boost to the 2-cm SCRC margin in adults with newly diagnosed malignant gliomas.

Systemic targeted radiation therapy and radioimmunotherapy continue to be important tools in the treatment of certain cancers. Because of their high energy and short path length, alpha particle emitters such as 211At are more effective than either external beam x- ray or in vivo beta radiation in delivering potentially curative doses of radiation. The limited clinical trials that have been conducted to date have yielded encouraging responses in some patients, e.g., malignant brain tumors. In order to escalate the additional necessary research and development in radiochemistry, radiobiology and efficacy evaluation of alpha particle radiotherapeutics, it is universally agreed that access to an affordable, reliable supply of 211At is warranted. In conjunction with the Department of Energy's intent to enhance stable and radioactive isotope availability for research applications, it is the primary objective of this project to improve 211At production and purification capabilities at Duke so that this radionuclide can be supplied to researchers at other institutions throughout the US.The most widely used 211At production method involves the α,2n reaction on Bismuth using a cyclotron with beams ≤ 28 MeV. Yields can be enhanced with use of an internal target that allows for a higher alpha fluence plus efficient heat dissipationmore » in the target. Both of these items are in place at Duke; however, in order to support production for multi-institutional use, irradiation campaigns in excess of 50 µAp and four hours duration will be needed. Further, post-irradiation processing equipment is lacking that will enable the distribution process. Financial support is sought for i) a shielded, ventilated processing/containment hood; ii) development of a post-irradiation target retrieval system; iii) fabrication of a 211At distillation and recovery module and iv) a performance review and, where needed, an enhancement of seven major subsystems that comprise the CS-30 Cyclotron. With these modifications in place, routine production of ≥200 mCi of At-211 should be readily achievable, given our methodological development of At-211 target preparation, internal target irradiation and dry distillation to recover the radionuclide.« less

Radiohalogenated agents are often the first line of pursuit in the development of new radiopharmaceuticals-whether antibodies, peptides, or small molecules-because of their ease of synthesis, lack of substantial steric perturbation of the original affinity agent (in some cases, providing enhanced affinity), and capacity to be transformed into therapeutics (in some cases, with a mere switch of an isotope). They often provide proof of a principle before optimization for pharmacokinetics or generation of radiometallated agents, when the latter are necessary. In particular, (18)F has been well integrated into normal clinical work flow in the form of (18)F-FDG for oncologic imaging, with reliable daily production and distribution to sites for immediate use, without the need for on-site preparation. Here we discuss radiohalogenated versions of imaging and therapeutic agents targeting the prostate-specific membrane antigen (PSMA); these were among the first such agents to be synthesized and used clinically. PSMA is highly expressed on prostate cancer epithelial cells and is currently being extensively investigated around the world as a target for imaging and therapy of prostate cancer. Additionally, the presence of PSMA on nonprostate tumor neovasculature has opened the possibility of PSMA-targeted molecules as generalizable cancer imaging and therapy agents. We focus on (18)F-labeled agents for PET, as they begin to redefine-along with the corresponding (68)Ga-labeled agents discussed elsewhere in this supplement to The Journal of Nuclear Medicine-the management of prostate cancer across a variety of clinical contexts.

alpha-Particle-emitting radionuclides, such as (211)At, with a 7.2-h half-life, may be optimally suited for the molecularly targeted radiotherapy of strategically sensitive tumor sites, such as those in the central nervous system. Because of the much shorter range and more potent cytotoxicity of alpha-particles than of beta-particles, (211)At-labeled agents may be ideal for the eradication of tumor cells remaining after surgical debulking of malignant brain tumors. The main goal of this study was to investigate the feasibility and safety of this approach in patients with recurrent malignant brain tumors.Chimeric antitenascin monoclonal antibody 81C6 (ch81C6) (10 mg) was labeled with 71-347 MBq of (211)At by use of N-succinimidyl 3-[(211)At]astatobenzoate. Eighteen patients were treated with (211)At-labeled ch81C6 ((211)At-ch81C6) administered into a surgically created resection cavity (SCRC) and then with salvage chemotherapy. Serial gamma-camera imaging and blood sampling over 24 h were performed.A total of 96.7% +/- 3.6% (mean +/- SD) of (211)At decays occurred in the SCRC, and the mean blood-pool percentage injected dose wasor = 0.3. No patient experienced dose-limiting toxicity, and the maximum tolerated dose was not identified. Six patients experienced grade 2 neurotoxicity within 6 wk of (211)At-ch81C6 administration; this neurotoxicity resolved fully in all but 1 patient. No toxicities of grade 3 or higher were attributable to the treatment. No patient required repeat surgery for radionecrosis. The median survival times for all patients, those with glioblastoma multiforme, and those with anaplastic astrocytoma or oligodendroglioma were 54, 52, and 116 wk, respectively.This study provides proof of concept for regional targeted radiotherapy with (211)At-labeled molecules in oncology. Specifically, the regional administration of (211)At-ch81C6 is feasible, safe, and associated with a promising antitumor benefit in patients with malignant central nervous system tumors.

Purpose To assess the efficacy and toxicity of intraresection cavity iodine-131–labeled murine antitenascin monoclonal antibody 81C6 (131I-m81C6) among recurrent malignant brain tumor patients. Patients and Methods In this phase II trial, 100 mCi of 131I-m81C6 was injected directly into the surgically created resection cavity (SCRC) of 43 patients with recurrent malignant glioma (glioblastoma multiforme [GBM], n = 33; anaplastic astrocytoma [AA], n = 6; anaplastic oligodendroglioma [AO], n = 2; gliosarcoma [GS], n = 1; and metastatic adenocarcinoma, n = 1) followed by chemotherapy. Results With a median follow-up of 172 weeks, 63% and 59% of patients with GBM/GS and AA/AO tumors were alive at 1 year. Median overall survival for patients with GBM/GS and AA/AO tumors was 64 and 99 weeks, respectively. Ten patients (23%) developed acute hematologic toxicity. Five patients (12%) developed acute reversible neurotoxicity. One patient (2%) developed irreversible neurotoxicity. No patients required reoperation for radionecrosis. Conclusion In this single-institution phase II study, administration of 100 mCi of 131I-m81C6 to recurrent malignant glioma patients followed by chemotherapy is associated with a median survival that is greater than that of historical controls treated with surgery plus iodine-125 brachytherapy. Furthermore, toxicity was acceptable. Administration of a fixed millicurie dose resulted in a wide range of absorbed radiation doses to the SCRC. We are now conducting a phase II trial, approved by the US Food and Drug Administration, using patient-specific 131I-m81C6 dosing, to deliver 44 Gy to the SCRC followed by standardized chemotherapy. A phase III multicenter trial with patient-specific dosing is planned.

Improving drug and macromolecular delivery of anti-cancer agents to tumours results in greater efficacy without increased toxicity. The current study was undertaken to assess the effects of the timing of injection of tumour specific and non-specific monoclonal antibodies (mAbs) relative to a hyperthermia treatment on tumour and normal tissue uptake. Using a local hyperthermia protocol of 45 min at 43 degrees C, uptake in tumour and normal tissues was measured at 1, 4, 12, 24, 48 and 72 h after injection. An anti-tenascin chimeric mAb, ch81C6, served as the specific mAb in a D-54 MG glioma xenograft mouse model. The chimeric mAb chTPS3.2 served as the control. A five-to-eight-fold increase in uptake of the tumour-targeted mAb was achieved in the heated tumours when compared with the non-heated tumours at 1 h. Differences in absolute tumour uptake of the specific mAb between the mice injected prior to hyperthermia and mice injected post-hyperthermia were seen only at 1 and 12 h. The median uptakes in the tumours of mice injected pre-heat were 25%ID/g at 1 h and 43.5%ID/g at 12 h, while in the animals injected post-hyperthermia the median uptakes were 45.5%ID/g and 80.2%ID/g, respectively. Blood levels of both the specific and non-specific mAbs were consistently higher over the initial 12 h period in the mice injected post-hyperthermia. Normal tissue uptake was also increased at most time points in the mice injected post-hyperthermia. The clinical importance of the differences in specific mAb uptake in tumour detected statistically at 1 and 12 h is questionable, given the highly variable nature of mAb uptake in vivo. Tumour targeting mAbs administered in combination with heat may be injected either prior to or immediately following hyperthermia treatment, with the expectation that levels of uptake in tumour will be relatively equivalent. Absolute normal tissue levels will be higher in patients receiving the mAb post-hyperthermia.

The utility of external beam radiotherapy for the treatment of malignant brain tumours is compromised by the need to avoid excessive radiation damage to normal CNS tissues. This review describes the current status of targeted radiotherapy, an alternative strategy for brain tumour treatment that offers the exciting prospect of increasing the specificity of tumour cell irradiation.

TP-38 is a recombinant chimeric targeted toxin composed of the EGFR binding ligand TGF-α and a genetically engineered form of the Pseudomonas exotoxin, PE-38. After in vitro and in vivo animal studies that showed specific activity and defined the maximum tolerated dose (MTD), we investigated this agent in a Phase I trial. The primary objective of this study was to define the MTD and dose limiting toxicity of TP-38 delivered by convection-enhanced delivery in patients with recurrent malignant brain tumors. Twenty patients were enrolled in the study and doses were escalated from 25ng/mL to 100 with a 40mL infusion volume delivered by two catheters. One patient developed Grade IV fatigue at the 100ng/mL dose, but the MTD has not been established. The overall median survival after TP-38 for all patients was 23 weeks whereas for those without radiographic evidence of residual disease at the time of therapy, the median survival was 31.9 weeks. Overall, 3 of 15 patients, with residual disease at the time of therapy, have demonstrated radiographic responses and one patient with a complete response and has survived greater than 83 weeks.

p-[18F]Fluorobenzylguanidine ([18F]PFBG) is a norepinephrine analog that has been developed as a positron emission tomography (PET) imaging radiopharmaceutical. Myocardial sympathetic innervation, neuroendocrine structures, and tumors can be noninvasively imaged with [18F]PFBG. In this study, the uptake characteristics of [18F]PFBG were investigated in 2 dogs with a spontaneous pheochromocytoma. The extent of the pheochromocytoma was well documented in both dogs on the PET study. The standardized uptake values within the pheochromocytomas were greater than 25 by 10 min, and were 37 and 50 by 45 min in each dog. A third dog that was suspected to have an adrenal mass was also studied. In this dog, the [18F]PFBG study was normal. Surgical exploration and adrenal biopsy confirmed the [15F]PFBG imaging findings in both dogs. In each dog, there was rapid blood-pool clearance (within 10 min after intravenous administration of the [18F]PFBG), with high uptake specific within the myocardium and adrenal medulla. The results indicate that [18F]PFBG may be useful for imaging canine pheochromocytomas and aid in differentiating adrenal masses.

Radionuclides conjugated to molecules that bind specifically to cancer cells are of great interest as a means to increase the specificity of radiotherapy. Currently, the methods to disseminate these targeted radiotherapeutics have been either systemic delivery or by bolus injection into the tumor or tumor resection cavity. Herein we model a potentially more efficient method of delivery, namely pressure-driven fluid flow, called convection-enhanced delivery (CED), where a device infuses the molecules in solution (or suspension) directly into the tissue of interest. In particular, we focus on the setting of primary brain cancer after debulking surgery, where the tissue margins surrounding the surgical resection cavity are infiltrated with tumor cells and the most frequent sites of tumor recurrence. We develop the combination of fluid flow, chemical kinetics, and radiation dose models needed to examine such protocols. We focus on Auger electron-emitting radionuclides (e.g. 67Ga, 77Br, 111In, 125I, 123I, 193mPt, 195mPt) whose short range makes them ideal for targeted therapy in this setting of small foci of tumor spread within normal tissue. By solving these model equations, we confirm that a CED protocol is promising in allowing sufficient absorbed dose to destroy cancer cells with minimal absorbed dose to normal cells at clinically feasible activity levels. We also show that Auger emitters are ideal for this purpose while the longer range alpha particle emitters fail to meet criteria for effective therapy (as neither would energetic beta particle emitters). The model is used with simplified assumptions on the geometry and homogeneity of brain tissue to allow semi-analytic solutions to be displayed, and with the purpose of a first examination of this new delivery protocol proposed for radionuclide therapy. However, we emphasize that it is immediately extensible to personalized therapy treatment planning as we have previously shown for conventional CED, at the price of requiring a fully numerical computerized approach.

Even with novel chemotherapeutic agents and external beam radiation therapy, the prognosis of neoplastic meningitis secondary to malignant melanoma is still dismal. The authors report a case study of a 46-year-old white female who presented with progressive hearing loss, severe headaches, nausea, vomiting, and a rapid decline in neurologic status. She was referred to Duke University Medical Center after conventional chemotherapy for malignant melanoma failed. She was enrolled in a Phase I trial of 131I-labeled monoclonal antibody Mel-14 F(ab′)2 fragment administered intrathecally. Within a year after her treatment, she recovered, having a normal neurologic exam except for residual bilateral hearing loss. The authors discuss dosimetry, preclinical, and clinical studies conducted with Mel-14 F(ab′)2 and introduce a potentially promising therapy option in the treatment of neoplastic meningitis in patients with malignant melanoma. Currently, the patient remains neurologically normal except for a mild bilateral hearing loss more than 4 years after treatment and has no radiographic evidence of neoplastic meningitis. Cancer 2001;91:1809–13. © 2001 American Cancer Society.

PURPOSE: To determine the maximum-tolerated dose (MTD) of iodine-131 (131I)–labeled 81C6 antitenascin monoclonal antibody (mAb) administered clinically into surgically created resection cavities (SCRCs) in malignant glioma patients and to identify any objective responses with this treatment. PATIENTS AND METHODS: In this phase I trial, newly diagnosed patients with malignant gliomas with no prior external-beam therapy or chemotherapy were treated with a single injection of 131I-labeled 81C6 through a Rickham reservoir into the resection cavity. The initial dose was 20 mCi and escalation was in 20-mCi increments. Patients were observed for toxicity and response until death or for a minimum of 1 year after treatment. RESULTS: We treated 42 patients with 131I-labeled 81C6 mAb in administered doses up to 180 mCi. Dose-limiting toxicity was observed at doses greater than 120 mCi and consisted of delayed neurotoxicity. None of the patients developed major hematologic toxicity. Median survival for patients with glioblastoma multiforme and for all patients was 69 and 79 weeks, respectively. CONCLUSION: The MTD for administration of 131I-labeled 81C6 into the SCRC of newly diagnosed patients with no prior radiation therapy or chemotherapy was 120 mCi. Dose-limiting toxicity was delayed neurologic toxicity. We are encouraged by the survival and toxicity and by the low 2.5% prevalence of debulking surgery for symptomatic radiation necrosis.

In this issue of Clinical Cancer Research , Borjesson et al. ([1][1]) report the results of the first clinical trial using the long-lived positron emitter 89Zr for immuno–positron emission tomography (PET). A series of 20 patients with squamous cell carcinoma of the head and neck were imaged up

A paired-label study was performed in athymic mice bearing subcutaneous D-54 MG human glioma xenografts to compare the localization of human/mouse anti-tenascin chimeric antibody 81C6 labeled by reaction with N-succinimidyl 3-[211At]astatobenzoate and N-succinimidyl 3-[131I]iodobenzoate. Over the 48-h observation period, the distribution of 211At- and 131I-labeled antibody were quite similar in tumor and normal tissues except stomach. These data were used to calculate human radiation doses for both intravenously and intrathecal administered 211At-labeled chimeric 81C6 using a quality factor of 5 for alpha-emissions.

Solid tumours have an elevated interstitial fluid pressure (IFP) due to the lack of normal lymphatics, increased permeability of tumour vasculature and an expanding cell population within a potentially limited space. This elevated IFP has been proposed to be an important barrier to the delivery of drugs and marcromolecules. We have demonstrated that local hyperthermia (4 h, 41.8 degrees C) is capable of significantly enhancing the uptake of radiolabelled monoclonal antibodies (mAbs) in D-54 MG glioma xenografts grown subcutaneously in athymic mice. To determine if this increased uptake was attributable to alterations in the tumour IFP, pressure measurements using the wick-in-needle technique were made in tumours after hyperthermia treatment. These pressure measurements were taken at various time points from 4 to 90 h following the initiation of the hyperthermia and compared with pressures taken concurrently in untreated tumours. In addition, pressures were measured following a 2 h, 41.8 degrees C hyperthermia treatment, a protocol which does not result in elevated uptake of radiolabeled mAbs. No significant differences were seen at any time point in IFP measured in the tumours receiving either hyperthermia treatment when compared with untreated tumours. Thus, we conclude that the mechanism by which this hyperthermia regimen enhances mAb uptake in this human glioma xenograft model is not due to alternations in tumour IFP.

Radionuclides such as 211At and 212Bi which decay by the emission of alpha-particles are attractive for certain applications of targeted radiotherapy. The tissue penetration of 212Bi and 211At alpha-particles is equivalent to only a few cell diameters, offering the possibility of combining cell-specific targeting with radiation of similar range. Unlike the beta-particles emitted by radionuclides such as 131I and 90Y, alpha-particles are radiation of high linear energy transfer and thus greater biological effectiveness. Several approaches have been explored for targeted radiotherapy with 212Bi- and 211At-labelled substances including colloids, monoclonal antibodies, metabolic precursors, receptor-avid ligands and other lower molecular weight molecules. An additional agent which exemplifies the promise of alpha-emitting radiopharmaceuticals is meta-[211At]astatobenzylguanidine. The toxicity of this compound under single-cell conditions, determined both by [3H]thymidine incorporation and by limiting dilution clonogenic assays, for human neuroblastoma cells is of the order of 1000 times higher than that of meta-[131I] iodobenzylguanidine. For meta-[211At] astatobenzylguanidine, the Do value was equivalent to only 6-7 211At atoms bound per cell. These results suggest that meta-[211At] astatobenzylguanidine might be valuable for the targeted radiotherapy of micrometastatic neuroblastomas.

A paired-label biodistribution was performed in athymic mice bearing SK-N-SH human neuroblastoma xenografts to compare the tissue uptake of meta-[211At]astatobenzylguanidine ([211At]MABG) and [131I]MIBG. Significantly higher (p < 0.05) uptake of [211At]MABG was seen in tumor (3.8 +/- 0.8% ID/g vs. 3.1 +/- 0.7% ID/g at 8 h) compared to [131I]MIBG. Desipramine reduced tumor uptake of [211At] MABG by 43%, suggesting that its accumulation was related to the specific uptake-1 mechanism. Higher uptake of [211At]MABG was also seen in normal tissue targets such as heart (6.0 +/- 0.9% ID/g vs. 4.5 +/- 0.8% ID/g at 8 h; p < 0.05). Pretreatment of mice with unlabeled MIBG increased tumor uptake of [211At]MABG by 1.5-fold while reducing uptake in heart and several other normal tissues. The vesicular uptake inhibitor tetrabenazine reduced heart uptake by 30% without reducing the tumor uptake. These results suggest such strategies might be useful for improving [211At]MABG tumor-to-normal tissue ratios.

The present study undertook to establish the dose (LD) of systematically administered (via tail vein) sodium [211At]astatide that would kill 10% (LD10) of exposed animals in two mouse models and to evaluate the resulting histologic lesions.Three dose escalation experiments were carried out using groups of 10 3- to 4-week-old, 20 +/- 2 g B6C3F1 mice, and one dose escalation experiment was carried out with groups of 10 4- to 6-week-old, 22 +/- 2 g BALB/c (nu/nu) mice. All animals were weighed daily and checked twice daily for general health; autopsies were performed within 12 h of death.The LD10 (95% confidence interval) level of free [211At]astatide at 360 days was 15.1 microCi (5.2-19.1 microCi) in B6C3F1 mice and was associated with a 37.8% weight difference from saline controls (p0.001). In the BALB/c (nu/nu) mice, the LD10 at 360 days was 7.7 microCi (0-14.2 microCi), while a dose of 10 microCi (0.42 microCi g(-1)) was associated with a 9.44% weight difference vs. saline controls (p0.05). Exclusive of the well-known effects on thyroid, [211At]astatide activity levels were associated with severe bone marrow depression, testicular atrophy, focal alopecia, and nuclear atypia of the epidermoid mucosa of the fore-stomach in the B6C3F1 mice; at activity levels approximating LD10 at 360 days, mild changes in the heart, liver, stomach, and spleen were observed. For BALB/c (nu/nu) mice, administration of 10 microCi was associated at autopsy with mild histologic lesions in the heart, stomach, liver, and spleen.These studies provide a basis for the design of further investigations of [211At]-labeled compounds as therapeutic agents.

The kinetics of para-[18F]fluorobenzylguanidine ([18F]PFBG) were investigated in a canine coronary artery occlusion model. Five dogs were imaged by positron emission tomography (PET) before and after complete surgical ligation of the left anterior descending coronary artery. PET studies included a 10-minute dynamic [13N]NH3 perfusion scan, followed 1 hour later by 3-hour dynamic [18F]PFBG scanning. [18F]PFBG and [13N]NH3 images demonstrated homogeneous myocardial uptake/perfusion before infarction. One hundred eighty minutes after [18F]PFBG administration, myocardial accumulation was decreased by 60% (day, 2, 0.0065% ±0.0015% injected dose/ml) and 58% (day 16, 0.0069%±0.003% injected dose/ml) compared with a similar myocardial region of interest from the preinfarction (0.016%±0.005% injected dose/ml) study. Myocardial accumulation of [13N]NH3 at 9 minutes showed a 52% (day 2) and 7% (day 16) decrease compared with the preinfarction study. The accumulation of [18F]PFBG in the infarction was decreased significantly at 120 and 180 minutes on all postinfarction studies (p=0.01). In three dogs a significant decrease in the myocardial norepinephrine concentration was documented in the area of infarction (237±94 ng/gm) versus the noninfarcted (1018±48 ng/gm) myocardium (p=0.001). A decreased accumulation of [18F]PFBG was observed in the area of myocardial infarct in this canine model. The magnitude of the decrease in [18F]PFBG was larger than that seen with [13N]NH3 on day 16 after infarction suggesting reperfusion and persistent sympathetic neuronal dysfunction.

Radionuclides which decay by the emission of alpha-particles are attractive for certain radioimmunotherapeutic applications. These include the treatment of lymphomas, compartmentally spread malignancies such as ovarian cancer and neoplastic meningitis, and micrometastatic disease. Two alpha-emitting radionuclides of interest for this purpose are 212Bi (60.6 min half life) and 211At (7.2 hr half life). Compared with the beta-emitters commonly used for radiotherapy, the alpha-particles of 212Bi and 211At are of higher energy, much shorter range (less than 100 microm), and considerably higher linear energy transfer. Preliminary results obtained in a variety of in vitro systems and in vivo models have documented the exquisite toxicity of alpha-particles and have established a basis for initiating radiotherapy trials in humans with monoclonal antibodies labeled with alpha-emitting radionuclides.

Malignant brain tumors are one of the most lethal cancer forms causing over 14,000 deaths each year. In spite of extensive research efforts over many decades, the median overall survival (OS) still remains at just 15 months for the common malignant giloma, glioblastoma multiforme (GBM). We have developed a multifunctional gold nanostar (GNS) probe that shows potential to improve cancer management by providing personalized cancer theranostics, a combination of diagnostic and therapeutic functions. The GNS nanoprobe has a tunable plasmon absorption peak in the near infrared (NIR) “tissue optical window” for deep-tissue penetration and tip-enhanced surface plasmon resonance, which makes the GNS as a superior nanoprobe for cancer detection using surface-enhanced Raman spectroscopy (SERS), two-photon photoluminescence (TPL) imaging and photothermal therapy with NIR light. We have also functionalized the GNS with iodine-124 radioisotopes for sensitive 3D positron emission tomography (PET) and achieved a 10-picomolar limit of detection (LOD). The GNS nanoprobe accumulates selectively in brain tumor due to the enhanced permeability and retention (EPR) effect, which is caused by defective endothelial tight junctions in blood vessels surrounding the tumor. On the contrary, the GNS nanoprobe has minimum uptake in normal brain tissue due to the intact blood-brain-barrier (BBB). The GNS uptake ratio of tumor-to-normal (T/N) brain tissue is measured to be up to 30:1 by using inductively coupled plasma mass spectrometry (ICP-MS). By combining the high detection sensitivity of PET imaging and specific accumulation of GNS in brain tumor, we have demonstrated the feasibility of 1-mm brain tumor detection with the I-124 labeled GNS nano radiotracer. The obtained T/N ratio is up to 10:1 with our GNS nanoprobe labeled with I-124, which is much better than that (typically less than 1.5:1) for traditional F-18 fluorodeoxyglucose (FDG) PET contrast agent due to high glucose uptake in normal brain tissue. Furthermore, we have utilized GNS's high TPL intensity to delineate brain tumor margin with excellent sensitivity and high spatial resolution (μm level). In addition to brain tumor imaging, we have successfully demonstrated photothermal ablation with a NIR laser taking advantage of GNS's superior photothermal conversion efficiency. We have also performed a 6-month toxicity study for the GNS nanoprobe and experiment results, including body weight monitoring, blood chemistry test, and H&E histopathology examination, show no deleterious effect after infusion of GNS nanoprobe up to 100 mg/kg dose. In summary, the GNS nanoprobe, with limited toxicity and multifunctional capabilities, exhibits great promise to fulfill the unmet needs in brain tumor management in future translational medicine investigations. Citation Format: Yang Liu, Austin Carpenter, Hsiangkuo Yuan, Zhengyuan Zhou, Michael Zalutsky, Ganesan Vaidyanathan, Hai Yan, Tuan Vo-Dinh. Gold nanostar as theranostic probe for brain tumor sensitive PET-optical imaging and image-guided specific photothermal therapy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4213.

In vitro and in vivo neuroblastoma models were used to determine whether improvements in tumour targeting in vivo and therapeutic efficacy in vitro could result from the use of no-carrier-added (n.c.a.) [131I]MIBG. Results were compared with use of the conventional therapy MIBG preparation (ex. [131I]MIBG) of lower specific activity which is produced by iodide exchange reaction. The efficacy of n.c.a. [131I]MIBG was compared with that of [131I]MIBG over a range of specific activities by the assessment of neuroblastoma spheroid growth delay. Whereas n.c.a. [131I]MIBG at a radioactivity concentration of 2 MBq/ml prevented the regrowth of 84% of spheroids, toxicity was significantly reduced by the addition of non-radiolabelled MIBG to the incubation medium. The time-dependent biodistribution of n.c.a. [131I]MIBG in nude mice bearing human neuroblastoma xenografts was compared with that of the conventional therapy radiopharmaceutical. The n.c.a. agent gave improved tumour uptake but also significantly greater accumulation in normal tissues known to accumulate MIBG such as heart, adrenal and skin. However, uptake and retention in the blood was unaltered. For all tissues examined, the 3-day calculations were undertaken to predict organ to tumour dose ratios which would result in human neuroblastoma patients with each of the [131I]MIBG preparations. These results suggest that significant therapeutic gain may be achieved by the use of n.c.a. [131I]MIBG as a treatment agent in neuroblastoma. neuroblastoma.

Brachytherapy is a common clinical technique involving implantation of sealed radioactive “seeds” within a tumor to selectively irradiate the tumor mass while minimizing systemic toxicity. To mitigate the disadvantages associated with complex surgical implantation and subsequent device removal procedures, we have developed an alternative approach using a genetically encoded peptide polymer solution composed of a thermally responsive elastin-like polypeptide (ELP) radiolabeled with 131I that self-assembles into radionuclide seeds upon intratumoral injection. The formation of these nontoxic and biodegradable polymer seeds led to prolonged intratumoral retention (∼85% ID/tumor 7 days postinjection) of the radionuclide, elicited a tumor growth delay in 100% of the tumors in two human xenografts (FaDu and PC-3), and cured more than 67% of tumor-bearing animals after a single administration of labeled ELP. These results suggest that in situ self-assembly of biodegradable and injectable radionuclide-containing polypeptide seeds could be a promising therapeutic alternative to conventional brachytherapy. Cancer Res; 72(22); 5956–65. ©2012 AACR.

Background This study evaluates the potential utility of a modular nanotransporter (MNT) for enhancing the nuclear delivery and cytotoxicity of the Auger electron emitter 125I in cancer cells that overexpress the epidermal growth factor receptor (EGFR). Methods MNTs are recombinant multifunctional polypeptides that we have developed for achieving selective delivery of short-range therapeutics into cancer cells. MNTs contain functional modules for receptor binding, internalization, endosomal escape and nuclear translocation, thereby facilitating the transport of drugs from the cell surface to the nucleus. The MNT described herein utilized EGF as the targeting ligand and was labeled with 125I using N-succinimidyl-4-guanidinomethyl-3-[125I]iodobenzoate (SGMIB). Membrane binding, intracellular and nuclear accumulation kinetics, and clonogenic survival assays were performed using the EGFR-expressing A431 epidermoid carcinoma and D247 MG glioma cell lines. Results [125I]SGMIB-MNT bound to A431 and D247 MG cells with an affinity comparable to that of native EGF. More than 60% of internalized [125I]SGMIB-MNT radioactivity accumulated in the cell nuclei after a 1-h incubation. The cytotoxic effectiveness of [125I]SGMIB-MNT compared with 125I-labeled bovine serum albumin control was enhanced by a factor of 60 for D247 MG cells and more than 1,000-fold for A431 cells, which express higher levels of EGFR. Conclusions MNT can be utilized to deliver 125I into the nuclei of cancer cells overexpressing EGFR, significantly enhancing cytotoxicity. Further evaluation of [125I]SGMIB-MNT as a targeted radiotherapeutic for EGFR-expressing cancer cells appears warranted.

Meta-iodobenzylguanidine (MIBG) is a drug which is selectively accumulated by the uptake-1 process in adrenergic tissues. When labelled with 131I, it may be used for the targetted radiotherapy of tumours such as phaeochromocytoma and neuroblastoma. This paper describes the preparation of carrier-free 131I-MIBG by radioiodination of meta-diazobenzylguanidine, and compares this process with one involving iododesilylation of meta-trimethylsilylbenzylguanidine. Both processes result in the formation of carrier-free 131I-MIBG whose specific activity at greater than 3 x 10(16) Bq mol-1 is at least 100 times higher than that of commercially available 131I-MIBG for therapeutic use. The therapeutic use of 131I-MIBG with a higher than usual specific activity is predicted to result in a greater target-to-nontarget ratio, and therefore enhanced efficacy because of an increased therapeutic index. As the radiochemical yield of the process involving the metadiazobenzylguanidine intermediate is only 13%, compared with 98% for the iododesilylation reaction, the latter is the preferred synthetic route.

The objective of this investigation was to evaluate small-field-of-view, ultra-high-resolution pinhole collimation for a rotating-camera SPECT system that could be used to image small laboratory animals. Pinhole collimation offers distinct advantages over conventional parallel-hole collimation when used to image small objects. Since geometric sensitivity increases markedly for points close to the pinhole, small-diameter and high-magnification pinhole geometries may be useful for selected imaging tasks when used with large-field-of-view scintillation cameras. The use of large magnifications can minimize the loss of system resolution caused by the intrinsic resolution of the scintillation camera. A pinhole collimator has been designed and built that can be mounted on one of the scintillation cameras of a triple-head SPECT system. Three pinhole inserts with approximate aperture diameters of 0.6, 1.2 and 2.0 mm have been built and can be mounted individually on the collimator housing. When a ramp filter is used with a three-dimensional (3D) filtered backprojection (FBP) algorithm, the three apertures have in-plane SPECT spatial resolutions (FWHM) at 4 cm of 1.5, 1.9 and 2.8 mm, respectively. In-air point source sensitivities at 4 cm from the apertures are 0.9, 2.6 and 5.7 counts s(-1) microCi(-1) (24, 70 and 154 counts s(-1) MBq(-1)) for the 0.6, 1.2 and 2.0 mm apertures, respectively. In vitro image quality was evaluated with a micro-cold-rod phantom and a micro-Defrise phantom using both the 3D FBP algorithm and a 3D maximum likelihood-expectation maximization (ML-EM) algorithm. In vivo image quality was evaluated using two (315 and 325 g) rats. Ultra-high-resolution pinhole SPECT is an inexpensive and simple approach for imaging small animals that can be used with existing rotating-camera SPECT system.

The authors have investigated standard SPECT techniques (rotating gamma cameras, multi-hole collimators, and filtered backprojection reconstruction) for imaging astatine-211 distributions. Since 211At emits alpha particles, this nuclide has potential for use in radiotherapy. The capability of imaging this nuclide would allow in vivo evaluation of the distribution and stability of potential 211At-labelled radiotherapeutic agents. 211At decay yields X-rays in the 77-92 keV range in addition to 500-900 keV gamma rays. The study evaluates the feasibility of SPECT imaging using the X-ray emissions of 211At. The authors have evaluated several collimators, with the determination that the medium-energy collimators are suitable, with 7% penetration (uncollimated counts versus collimated counts). Several phantoms were imaged and attenuation coefficients were measured (narrow-beam mu =0.182 cm-1 for 77-80 keV X-rays in water).

Targeted radiotherapy using agents tagged with α-emitting radionuclides is gaining traction with several clinical trials already undertaken or ongoing, and others in the advanced planning stage. The most commonly used α-emitting radionuclides are 213Bi, 211At, 223Ra and 225Ac. While each one of these has pros and cons, it can be argued that 211At probably is the most versatile based on its half life, decay scheme and chemistry. On the other hand, for targeting bone metastases, 223Ra is the ideal radionuclide because simple cationic radium can be used for this purpose. In this review, we will discuss the recent developments taken place in the application of 211At-labeled radiopharmaceuticals and give an overview of the current status of 223Ra for targeted α-particle radiotherapy.

Introduction Carcinomatous meningitis (CM) is a devastating disease characterized by the dissemination of malignant tumor cells into the subarachnoid space along the brain and spine. Systemic treatment with monoclonal antibody (mAb) trastuzumab can be effective against HER2-positive systemic breast carcinoma but, like other therapies, is ineffective against CM. The goal of this study was to evaluate the therapeutic effect of α-particle emitting 211 At-labeled trastuzumab following intrathecal administration in a rat model of breast carcinoma CM. Methods Athymic rats were injected intrathecally with MCF-7/HER2-18 breast carcinoma cells through a surgically implanted indwelling intrathecal catheter. In Experiment 1, animals received 33 or 66 μCi 211 At-labeled trastuzumab, cold trastuzumab or saline. In Experiment 2, animals were inoculated with a lower tumor burden and received 46 or 92 μCi 211 At-labeled trastuzumab or saline. In Experiment 3, animals received 28 μCi 211 At-labeled trastuzumab, 30 μCi 211 At-labeled TPS3.2 control mAb or saline. Histopathological analysis of the neuroaxis was performed at the end of the study. Results In Experiment 1, median survival increased from 21 days for the saline and cold trastuzumab groups to 45 and 48 days for 33 and 66 μCi 211 At-labeled trastuzumab, respectively. In Experiment 2, median survival increased from 23 days for saline controls to 68 and 92 days for 46 and 92 μCi 211 At-labeled trastuzumab, respectively. In Experiment 3, median survival increased from 20 days to 29 and 36 days for animals treated with 211 At-labeled TPS3.2 and 211 At-labeled trastuzumab, respectively. Long-term survivors were observed exclusively in the 211 At-trastuzumab-treated groups. Conclusion Intrathecal 211 At-labeled trastuzumab shows promise as a treatment for patients with HER2-positive breast CM.

Purpose: To generate and evaluate a modular recombinant transporter (MRT) for targeting {sup 211}At to cancer cells overexpressing the epidermal growth factor receptor (EGFR). Methods and Materials: The MRT was produced with four functional modules: (1) human epidermal growth factor as the internalizable ligand, (2) the optimized nuclear localization sequence of simian vacuolating virus 40 (SV40) large T-antigen, (3) a translocation domain of diphtheria toxin as an endosomolytic module, and (4) the Escherichia coli hemoglobin-like protein (HMP) as a carrier module. MRT was labeled using N-succinimidyl 3-[{sup 211}At]astato-5-guanidinomethylbenzoate (SAGMB), its {sup 125}I analogue SGMIB, or with {sup 131}I using Iodogen. Binding, internalization, and clonogenic assays were performed with EGFR-expressing A431, D247 MG, and U87MG.wtEGFR human cancer cell lines. Results: The affinity of SGMIB-MRT binding to A431 cells, determined by Scatchard analysis, was 22 nM, comparable to that measured before labeling. The binding of SGMIB-MRT and its internalization by A431 cancer cells was 96% and 99% EGFR specific, respectively. Paired label assays demonstrated that compared with Iodogen-labeled MRT, SGMIB-MRT and SAGMB-MRT exhibited more than threefold greater peak levels and durations of intracellular retention of activity. SAGMB-MRT was 10-20 times more cytotoxic than [{sup 211}At]astatide for all three cell lines. Conclusion: Themore » results of this study have demonstrated the initial proof of principle for the MRT approach for designing targeted {alpha}-particle emitting radiotherapeutic agents. The high cytotoxicity of SAGMB-MRT for cancer cells overexpressing EGFR suggests that this {sup 211}At-labeled conjugate has promise for the treatment of malignancies, such as glioma, which overexpress this receptor.« less

The prognosis for patients with malignant primary or metastatic brain tumors remains poor.1 Failure of conventional therapies can be attributed, at least in part, to their lack of specificity for neoplastic tissue, which results in dose-limiting systemic or neurologic toxicity.2 Protein cytotoxins produced by bacteria and plants represent a novel class of therapeutic agents that can be conjugated to specific ligands and delivered regionally to target tumors more specifically.3–6 These targeted toxins have produced dramatic clinical responses in therapeutically refractory patients with leukemia7–9 and lymphoma.10,11 The efficacy of these agents is likely related to their potency, specificity, immunity to resistance mechanisms, and lack of dependence on cell-cycle kinetics. Results in solid tumors, however, have been less encouraging, possibly because these large molecules have difficulty penetrating solid tumors.12–15 The surface expression of targeted antigens may also be lower and the heterogeneity of antigen expression may be greater in solid neoplasms; as a result, a single antibody is less likely to target all neoplastic cells. Delivery of high-molecular-weight agents, such as targeted toxins, to tumors within the intracerebral compartment is even more challenging because these tumors are further isolated from the systemic circulation by the restrictive blood-brain barrier (BBB).13,14,16–19 Regional intracerebral drug delivery, however, bypasses the BBB and has the potential to deliver high concentrations of the therapeutic agent directly to the site of the tumor, thereby reducing the risk of systemic toxicity. The innovative regional drug delivery technique of convection-enhanced delivery (CED) uses a fluid pressure gradient to infuse therapeutic molecules directly into the interstitial spaces of the brain that are infiltrated with tumor.20–23 The potential of this simple approach has been clearly demonstrated in preclinical studies by others, especially Lonser and Oldfield’s group, and more recently by ourselves.19,20,22–37 The results of early clinical studies21,38–47 have also shown some potential efficacy, but they have not attempted to assess the adequacy of drug distribution in humans. TP-38 is a 43.5-kDa recombinant chimeric protein containing a genetically engineered form of the cytotoxic Pseudomonas exotoxin (PE). By replacing the native binding domain of PE with transforming growth factor-α (TGF-α), the TP-38 construct has been specifically targeted to the epidermal growth factor receptor (EGFR). EGFR is frequently overexpressed in malignant gliomas and metastatic tumors of the brain,48 but is expressed at only very low levels within normal human brain.49,50 In this phase I study we sought to determine the maximum tolerated dose (MTD), dose-limiting toxicity (DLT), and intracerebral distribution of TP-38 using the intracerebral infusion technique of CED. We show that CED of TP-38 is well tolerated at effective doses and provides some encouraging radiographic responses. However, we also show that the potential efficacy of agents delivered by CED may be severely constrained by inconsistent and ineffective infusion in many patients.

Adults with primary malignant glioma have an unacceptably poor outcome. Most of these tumors recur at or adjacent to the site of origin, which indicates that failure to eradicate local tumor growth is a major factor contributing to poor outcome. Therefore, locoregional therapies may improve local control and overall outcome for malignant glioma patients. Malignant gliomas selectively express several factors that are not present on normal CNS tissue. Regional administration of radiolabeled monoclonal antibodies targeting tumor-specific antigens expressed by malignant gliomas offers an innovative therapeutic strategy that has recently demonstrated encouraging antitumor activity and acceptable toxicity in clinical trials at a number of centers. Most studies have utilized monoclonal antibodies against tenascin-C, an extracellular matrix glycoprotein ubiquitously expressed by malignant gliomas. This review summarizes clinical trials performed using radiolabeled antitenascin-C monoclonal antibodies for malignant glioma patients to date and highlights future plans to further develop this therapeutic strategy.

An attractive feature of targeted radionuclide therapy is the ability to select radionuclides and targeting vehicles with characteristics that are best suited for a particular clinical application. One combination that has been receiving increasing attention is the use of monoclonal antibodies (mAbs) specifically reactive to receptors and antigens that are expressed in tumor cells to selectively deliver the alpha-particle-emitting radiohalogen astatine-211 (211At) to malignant cell populations. Promising results have been obtained in preclinical models with multiple 211At-labeled mAbs; however, translation of the concept to the clinic has been slow. Impediments to this process include limited radionuclide availability, the need for suitable radiochemistry methods operant at high activity levels and lack of data concerning the toxicity of alpha-particle emitters in humans. Nonetheless, two clinical trials have been initiated to date with 211At-labeled mAbs, and others are planned for the near future.

The intratumoral delivery of radioactive therapeutics, a clinical technique known as brachytherapy, has proven very successful in treating solid cancers and become a prevalent option for treating prostate, breast, and melanoma tumors. The method typically involves the permanent implantation of 80-120 titanium ‘seeds’ encapsulating radioactive isotopes and requires expert spatial placement to ensure full dose coverage. The disadvantages of the seeds include a complicated implantation process, the permanence of the implants, migration of seeds to off-target sites, and that the titanium shielding limits the application of radionuclides which could treat more aggressive forms of cancer. As such, we have developed a next-generation brachytherapy alternative to solid seeds using an injectable, biodegradable, biocompatible and thermosensitive elastin-like polypeptide (ELP). Through recombinant design and synthesis, ELP exhibits a thermal phase transition (soluble-to-insoluble) when heated above 21°C. This allows radionuclide-bearing ELP to be easily handled as a fluid at room temperature, but enables spontaneous self-assembly into a stable brachytherapy gel upon intratumoral injection at body temperature. Previously, we demonstrated the efficacy of ELP brachytherapy in treating both s.c. human hypopharyngeal tumors and orthotopic human prostate tumors xenografted in mice models. Treatment options for pancreatic cancer remain limited due to a large proportion of patients presenting with advanced disease at the time of diagnosis. Trials investigating the use of brachytherapy as a neo-adjuvant for shrinking pancreatic tumors prior to surgery or as an adjuvant to inhibit recurrence have provided encouragement. By conjugating the more potent radionuclide 131-Iodine to ELP, we investigated the ability of ELP brachytherapy to treat more clinically aggressive pancreatic tumors. Utilizing a human CRL-1687 cell line transfected to express luciferase (BxPc3-luc2), human pancreatic tumors were xenografted into an orthotopic, athymic nu/nu mouse model. Upon reaching 150 mm^3 in size, mice were treated either with [131I]-ELP brachytherapy (n = 6), at a dose of 5.83 μCi/mm^3, or with an ELP-only control injection (n = 6). Tumor response was tracked non-invasively using luminescent imaging for 60 days. The ELP brachytherapy demonstrated significant tumor inhibition (p < 0.05, one-way ANOVA) and no appreciable side effects as determined by weight loss. The therapy group experienced a significant survival benefit, with a median duration of 27 days (p < 0.005, log-rank test), when compared to control mice. These results, while promising as a proof of concept, demonstrate that further development is required to determine the both the appropriate radiation dose and to intratumoral coverage for translational application. Citation Format: Jeffrey Laurence Schaal, Wenge Liu, Ashutosh Chilkoti, Xinghai Li, Eric Mastria, Michael R. Zalutsky. Next-generation brachytherapy: a preclinical study of a thermally stabilized biopolymer gel for delivering intratumoral radionuclide therapy in a pancreatic tumor mouse model. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 1809. doi:10.1158/1538-7445.AM2015-1809

The neurohistological findings in patients treated with targeted beta emitters such as (131)I are poorly described. We report a histopathologic analysis from patients treated with combined external beam therapy and a brachytherapy consisting of a (131)I-labeled monoclonal antibody (mAb) injected into surgically created resection cavities during brain tumor resections.Directed tissue samples of the cavity walls were obtained because of suspected tumor recurrence from 28 patients. Samples and clinical follow-up were evaluated on all patients (Group A) based on total radiation dose received and a subset of these (n=18; Group B, proximal therapy subset) who had received external beam therapy withinor=3 months of mAb therapy and undergoing 26 biopsies over 37 months. Histologic outcomes were "proliferative glioma," "quiescent glioma" and negative for neoplasm. Statistical analysis was used to assess the casual relation between total absorbed dose ((131)I-mAb+external beam) and histologic diagnosis.The lesions observed after (131)I-mAb therapy were qualitatively similar to those reported for other types of radiation therapy; however, the high localized dose rate and absorbed doses produced by the short range of (131)I beta particles seem to have resulted in an earlier necrotic reaction in the tumor bed. Among all 28 (Group A) patients, median survival from tissue analysis after mAb therapy depended on histopathology and total radiation absorbed dose. Median survival for patients with tissue classified as proliferative glioma, quiescent glioma and negative for neoplasm were 3.5, 15 and 27.5 months, respectively. Without categorization, total dose was a significant predictor of survival (P.002) where patients with higher doses had better prognoses. For example, median survival in patients receiving a total radiation dose greater than 86 Gy was 19 months compared with 7 months for those receiving less than 86 Gy.Histopathologic analysis correlated with prognosis. Among all patients (Group A) there was a significant correlation between biopsy outcome, survival, and total radiation absorbed dose. Among the Group B proximal therapy patients, the neuropathologic changes were qualitatively similar to those described for external beam therapy and interstitial brachytherapy.