Joseph O’Donoghue

A preclinical model for noninvasive imaging of hypoxia-induced gene expression; comparison with an exogenous marker of tumor hypoxia.

PurposeHypoxia is associated with tumor aggressiveness and is an important cause of resistance to radiation therapy and chemotherapy. Assays of tumor hypoxia could provide selection tools for hypoxia-modifying treatments. The purpose of this study was to develop and characterize a rodent tumor model with a reporter gene construct that would be transactivated by the hypoxia-inducible molecular switch, i.e., the upregulation of HIF-1.MethodsThe reporter gene construct is the herpes simplex virus 1-thymidine kinase (HSV1-tk) fused with the enhanced green fluorescent protein (eGFP) under the regulation of an artificial hypoxia-responsive enhancer/promoter. In this model, tumor hypoxia would up-regulate HIF-1, and through the hypoxia-responsive promoter transactivate the HSV1-tkeGFP fusion gene. The expression of this reporter gene can be assessed with the 124I-labeled reporter substrate 2′-fluoro-2′-deoxy-1-β-d-arabinofuranosyl-5-iodouracil (124I-FIAU), which is phosphorylated by the HSV1-tk enzyme and trapped in the hypoxic cells. Animal positron emission tomography (microPET) and phosphor plate imaging (PPI) were used in this study to visualize the trapped 124I-FIAU, providing a distribution of the hypoxia-induced molecular events. The distribution of 124I-FIAU was also compared with that of an exogenous hypoxic cell marker, 18F-fluoromisonidazole (FMISO).ResultsOur results showed that 124I-FIAU microPET imaging of the hypoxia-induced reporter gene expression is feasible, and that the intratumoral distributions of 124I-FIAU and 18F-FMISO are similar. In tumor sections, detailed radioactivity distributions were obtained with PPI which also showed similarity between 124I-FIAU and 18F-FMISO.ConclusionThis reporter system is sufficiently sensitive to detect hypoxia-induced transcriptional activation by noninvasive imaging and might provide a valuable tool in studying tumor hypoxia and in validating existing and future exogenous markers for tumor hypoxia.

Hypoxia in microscopic tumors

Tumor hypoxia has been commonly observed in a broad spectrum of primary solid malignancies. Hypoxia is associated with tumor progression, increased aggressiveness, enhanced metastatic potential and poor prognosis. Hypoxic tumor cells are resistant to radiotherapy and some forms of chemotherapy. Using an animal model, we recently showed that microscopic tumors less than 1mm diameter were severely hypoxic. In this review, models and techniques for the study of hypoxia in microscopic tumors are discussed.

Pilot study of 68Ga-DOTA-F(ab′)2-trastuzumab in patients with breast cancer

Objective68Ga-1,4,7,10-Tetraazacyclododecane-N,N′,N′′,N′′′-tetraacetic acid (DOTA)-F(ab′)2-trastuzumab [68Ga-DOTA-F(ab′)2-trastuzumab] has been developed at our institution as a positron imaging reagent for assessing human epidermal growth factor receptor 2 (HER2) expression status by in-vivo imaging. Initial studies on animals demonstrated promising results in the monitoring of treatment response to heat shock protein 90-targeted drugs that inhibit the client protein HER2. We report here our initial clinical experience in the assessment of the toxicity, pharmacokinetics, biodistribution, and dosimetry profile of 68Ga-DOTA-F(ab′)2-trastuzumab with PET/computed tomography using a mean of 236 MBq/5 mg administered intravenously. Materials and methodsA group of 16 women with breast cancer were enrolled in this study. The one patient who did not receive 68Ga-DOTA-F(ab′)2-trastuzumab was excluded from analysis. Both HER2-negative (n=7) and HER2-positive (n=8) cases were studied. Among the latter, seven had undergone trastuzumab treatment previously and one had not. ResultsIt was determined that 68Ga-DOTA-F(ab′)2-trastuzumab was well tolerated, with a T½ of ∼3.6±0.9 h; the critical organ was the kidney, with a mean dose of 0.383 cGy/37 MBq; and tumor targeting was seen in 4/8 patients with HER2-positive disease. ConclusionThe reagent is safe, and assessments through additional studies in a better-defined group of patients, using larger administered masses of antibodies, with a better immunoreactive fraction are needed.

RADIATION SAFETY CONSIDERATIONS FOR THE USE OF 223RaCl2 DE IN MEN WITH CASTRATION-RESISTANT PROSTATE CANCER

AbstractThe majority of patients with late stage castration-resistant prostate cancer (CRPC) develop bone metastases that often result in significant bone pain. Therapeutic palliation strategies can delay or prevent skeletal complications and may prolong survival. An alpha-particle based therapy, radium-223 dichloride (223RaCl2), has been developed that delivers highly localized effects in target areas and likely reduces toxicity to adjacent healthy tissue, particularly bone marrow. Radiation safety aspects were evaluated for a single comprehensive cancer center clinical phase 1, open-label, single ascending-dose study for three cohorts at 50, 100, or 200 kBq kg−1 body weight. Ten patients received administrations, and six patients completed the study with 1 y follow-up. Dose rates from patients administered 223Ra dichloride were typically less than 2 &mgr;Sv h−1 MBq−1 on contact and averaged 0.02 &mgr;Sv h−1 MBq−1 at 1 m immediately following administration. Removal was primarily by fecal excretion, and whole body effective half-lives were highly dependent upon fecal compartment transfer, ranging from 2.5–11.4 d. Radium-223 is safe and straightforward to administer using conventional nuclear medicine equipment. For this clinical study, few radiation protection limitations were recommended post-therapy based on facility evaluations. Specific precautions are dependent on local regulatory authority guidance. Subsequent studies have demonstrated significantly improved overall survival and very low toxicity, suggesting that 223Ra may provide a new standard of care for patients with CRPC and bone metastases.

Indium 111-labeled J591 anti-PSMA antibody for vascular targeted imaging in progressive solid tumors

BackgroundJ591 is a monoclonal antibody that targets the external domain of the prostate-specific membrane antigen (PSMA). Besides prostate cancer cells, it also targets the neovasculature of non-prostate solid tumors. We provide an analysis of the antibody mass-dose dependency of lesion uptake and normal tissue retention, together with an assessment of lesion detectability using 111In-J591 imaging, compared with conventional imaging in patients with a variety of solid tumors.MethodsTwenty patients in six cohorts received fixed amounts (5, 10, 20, 40, 60, and 100 mg) of J591 in a phase I trial. A maximum of four administrations per patient was given, with each administration separated by 3 weeks. All antibody administrations included 370 MBq (10 mCi) of 111In labeled to 2 mg of J591 via the chelating agent DOTA. Three whole body (WB) gamma camera scans with at least one SPECT scan, along with multiple WB count-rate measurements and blood samples, were obtained for all patients. The effect of escalating antibody mass on lesion uptake and normal tissue retention was evaluated using lesion, liver, serum, and WB residence times and ratios thereof for each treatment cycle. Lesion detectability using 111In-J591 imaging was compared to the standard imaging on a lesion-by-lesion basis.ResultsA total of 170 lesions in 20 patients were detected by standard or 111In-J591 imaging. 111In-J591 targeted both skeletal and soft tissue diseases in all tumor types. 111In-J591 imaging identified 74% (20/27) of skeletal lesions, 53% (18/34) of nodes, and 64% (70/109) of other soft tissue/organ lesions. There was increasing 111In-J591 uptake in lesions with increasing antibody mass-dose, coupled with decreasing retention in the liver for increments up to 20 mg, and no significant change at higher antibody mass.ConclusionsRadiolabeled J591 antibody has potential as a targeting agent for solid tumor vasculature and lesion detection. Bone and soft tissue lesions arising from tumors of diverse origin were targeted by the anti-PSMA antibody J591. For the detection of lesions in these tumors by J591 antibody scans, an antibody mass of 20 mg is adequate. The optimal time of imaging is 5 to 7 days post-injection.

Pilot study of PET imaging of 124I-iodoazomycin galactopyranoside (IAZGP), a putative hypoxia imaging agent, in patients with colorectal cancer and head and neck cancer

BackgroundHypoxia within solid tumors confers radiation resistance and a poorer prognosis. 124I-iodoazomycin galactopyranoside (124I-IAZGP) has shown promise as a hypoxia radiotracer in animal models. We performed a clinical study to evaluate the safety, biodistribution, and imaging characteristics of 124I-IAZGP in patients with advanced colorectal cancer and head and neck cancer using serial positron emission tomography (PET) imaging.MethodsTen patients underwent serial whole-torso (head/neck to pelvis) PET imaging together with multiple whole-body counts and blood sampling. These data were used to generate absorbed dose estimates to normal tissues for 124I-IAZGP. Tumors were scored as either positive or negative for 124I-IAZGP uptake.ResultsThere were no clinical toxicities or adverse effects associated with 124I-IAZGP administration. Clearance from the whole body and blood was rapid, primarily via the urinary tract, with no focal uptake in any parenchymal organ. The tissues receiving the highest absorbed doses were the mucosal walls of the urinary bladder and the intestinal tract, in particular the lower large intestine. All 124I-IAZGP PET scans were interpreted as negative for tumor uptake.ConclusionsIt is safe to administer 124I-IAZGP to human subjects. However, there was insufficient tumor uptake to support a clinical role for 124I-IAZGP PET in colorectal cancer and head and neck cancer patients.Trial registrationClinicalTrials.gov NCT00588276

Phase I trial of zirconium 89 (Zr89) radiolabeled J591 in metastatic castration-resistant prostate cancer (mCRPC).

31 Background: Presently, there are no means of accurately and reproducibly imaging bone metastases for patients with mCRPC. We are investigating a prostate-specific imaging method that can directly visualize metastases using PSMA as a target and Zr89 radiolabeled anti-PSMA antibody J591 as a tracer. Following the FDA roadmap for biomarker development, we report the first human data on the tracer and its preliminary analytic validation. METHODS 5 mCi of Zr89-J591 was administered intravenously and 4 serial PET/CT scans were obtained within the following time intervals post-injection: 2-4 hours, 18-30 hours, 2-5 days, and 6-7 days. Pharmacokinetic (PK) sampling occurred at 5, 30, and 60 minutes post-injection, and at each PET scan. Patients underwent standard cross-sectional imaging, bone scintigraphy, and FDG PET scanning. Metastatic sites were biopsied in order of preference: Zr89-J591 and FDG positivity, then Zr89-J591 and FDG mismatch, and lastly standard imaging and any PET mismatch. RESULTS Ten patients were scanned, 132 Zr89-J591 positive sites were detected, and 12 lesions (5 bone, 6 node, 1 lung) in 8 patients were biopsied. Tissue correlation: 12/12 biopsies were positive for cancer. 11/12 biopsied lesions (5 bone, 5 node, 1 lung) were positive on Zr89-J591 imaging. 10/12 were positive by standard scans or FDG. 1 lesion was negative by Zr89-J591 PET, but positive by other modalities. Immunokinetics: Blood clearance T1/2α: 7 +/- 4.5 h (1.1-14 h); T1/2β: 62 +/- 13 h (51-89 h); Whole body clearance T1/2: 219 +/- 48 h (153-317 h). Dosimetry: Maximum retention in liver 8 +/- 1.5 SUV; Kidney 4.1 +/- 0.8 SUV; Tumor 8.2 +/- 6.5 SUV (mean SUV in bone 11.13 +/- 7.57 vs. soft tissue 4.31 +/- 2.59). Optimal time for patient imaging after injection, in terms of tumor to background ratios was 7 +/- 1 days. CONCLUSIONS Zr89-J591 imaging demonstrates excellent tumor localization, pathology correlation, and can demonstrate the presence of tumor even when lesions are negative by standard imaging or FDG PET imaging. PK properties of the tracer have been defined. Use of Zr89-J591 to direct biopsies of metastases provides excellent yield. Further studies to examine reproducibility and post-treatment effects are planned. CLINICAL TRIAL INFORMATION NCT01543659.

MO-D-I-609-08: Validation of PET Hypoxia Tracers by Autoradiography and Fluorescent Microscopy

Purpose: To develop a method for PET hypoxia tracer validation based on statistical analysis of the spatial correspondence between the intratumoral distributions of tracer uptake (assessed by digital autoradiography) and pimonidazole, an independent marker of hypoxia (assessed by immunofluorescent microscopy). The utility of the method was demonstrated by applying it to three PET hypoxia tracers, IAZGP, FMISO and Cu‐ATSM. Method and Materials: Eight rats bearing R3327‐AT tumors were divided into four groups of two animals each. Group♯1 was injected with 18F‐FMISO and sacrificed 2hr later. Group♯2 was injected with 124I‐IAZGP and sacrificed 3hr later. Groups♯3 and ♯4 were injected with 64Cu‐ATSM and sacrificed, respectively, 24hr or 1hr later. Pimonidazole was administered to all animals 2hr before sacrifice. Tumors were excised, frozen and sectioned. Digital autoradiograms of the tracer distribution were obtained from selected sections and co‐registered with images of pimonidazole‐associated immunofluorescence derived from adjacent sections. The statistical analysis of association between PET tracer uptake and pimonidazole immunofluorescent staining intensity was then performed on a pixel‐by‐pixel basis. Results: For rats from group♯1 correlation coefficients between pimonidazole‐associated immunofluorescent staining intensity and 18F‐FMISO uptake were 0.84 and 0.75. For group♯2 rats correlations between pimonidazole and 124I‐IAZGP were 0.85 and 0.77. For group♯3 (64Cu‐ATSM administered 24hr prior to sacrifice), the correlations with pimonidazole were 0.61 and 0.64. However, for group♯4 (64Cu‐ATSM administered 1hr prior to sacrifice), the correlation coefficients were −0.76 and −0.77, demonstrating that in this tumormodel64Cu‐ATSM uptake was not indicative of hypoxia at early times post injection. Conclusion: The proposed method enables evaluation of the degree of spatial correspondence between distributions of a PET tracer and alternative markers (either endogenous or exogenous) or tracers of the biological process under study. This method can be used for the in‐vivo validation of any nuclear medicine tracer.