Flavio Forrer

Glucagon-like peptide-1 receptor imaging for localization of insulinomas.

CONTEXT The surgical removal of insulinomas is hampered by difficulties to localize it using conventional radiological procedures. Recently these tumors were shown to exhibit a very high density of glucagon-like peptide-1 receptors (GLP-1R) in vitro that may be used as specific targets for in vivo receptor radiolabeling. OBJECTIVE The objective of the study was to test the 111In-labeled GLP-1R agonist 111In-DOTA-exendin-4 in localizing insulinomas using single photon emission computed tomography in combination with computed tomography images. DESIGN This was a prospective open-label investigation. SETTING The study was conducted at three tertiary referral centers in Switzerland. PATIENTS Patients included six consecutive patients with proven clinical and biochemical endogenous hyperinsulinemic hypoglycemia. INTERVENTION (111)In-DOTA-exendin-4 was administered iv at a dose of about 90 MBq (30 microg peptide) over 5 min. Whole-body planar images of the abdomen were performed at 20 min, 4 h, 23 h, 96 h, and up to 168 h after injection. After surgical removal of the insulinomas, GLP-1R expression was assessed in the tumor tissue in vitro by GLP-1R autoradiography. MAIN OUTCOME MEASURE The detection rate of insulinomas was measured. RESULTS In all six cases, the GLP-1R scans successfully detected the insulinomas identified using conventional methods in four cases. By using a gamma-probe intraoperatively, GLP-1R detection permitted a successful surgical removal of the tumors in all patients, diagnosed histopathologically as five pancreatic and one extrapancreatic insulinomas. In vitro GLP-1R autoradiography showed a high density of GLP-1R in all tested insulinomas. CONCLUSION In vivo GLP-1R imaging is an innovative, noninvasive diagnostic approach that successfully localizes small insulinomas pre- and intraoperatively and that may in the future affect the strategy of insulinoma localization.

Radiopeptide Imaging and Therapy in Europe

Receptor targeting with radiolabeled peptides has become an important topic, particularly in nuclear oncology. Strong research efforts are under way in radiopharmaceutical science laboratories and in nuclear medicine departments in Europe. The target receptors belong to the large family of G-protein–coupled receptors. The prototypes of these radiopeptides are based on analogs of somatostatin targeting somatostatin receptor–positive tumors, particularly well-differentiated neuroendocrine tumors. These radiopeptides have an important impact not only on diagnosis but also on targeted radionuclide therapy of these tumors. Besides the registered radiopeptide 111In-pentetreotide, efficient SPECT tracers labeled with 99mTc and PET agents based on generator-produced 68Ga have been developed and used in the clinic. In parallel to the development of diagnostic agents, radiopeptides labeled with the β− emitters 90Y and 177Lu are also widely used. Because the same chelators and therefore the same conjugates can be used in diagnosis and therapy, they constitute ideal theranostic pairs. This progress is driven not only by scientists and clinicians but also by patient interest groups. New radiopeptides targeting other G-protein–coupled receptors are entering the clinic, among them glucagon-like peptide 1 receptor–targeting molecules. This receptor is overexpressed on literally all benign insulinomas. 111In-labeled derivatives of the insulinotropic 39-mer peptide exendin-4 were beneficial in the pre- and perioperative localization of these benign lesions. In contrast, lack of localization may indicate malignant insulinoma. The bombesin- and gastrin-releasing peptide receptor family is potentially important because these receptors are overexpressed on major human tumors such as prostate tumors, breast tumors, gastrointestinal stromal tumors, and vessels of ovarian cancer. 99mTc-labeled peptides for SPECT and 68Ga-, as well as 64Cu-labeled agonists or antagonists, have been studied in breast tumors, prostate tumors, gastrointestinal stromal tumors, and gliomas with considerable success. A phase I therapeutic study with a 177Lu-labeled agonist has been completed. There are not enough clinical data available to reveal the significance of these new modalities in patient care, but several phase I studies are under way in larger patient cohorts using PET agents. Another G-protein–coupled receptor with high overexpression on human tumors is the gastrin/cholecystokinin-2 receptor. It is overexpressed in more than 90% of cases of medullary thyroid cancer, in small cell lung cancer, and in a subgroup of neuroendocrine tumors. Correlating with in vitro receptor localization using autoradiography of 27 patients with metastasized medullary thyroid cancer, SPECT or planar imaging of these patients resulted in a 95% sensitivity of tumor localization. Finally, another G-protein–coupled receptor is found in brain tumors and peritumoral vessels. Literally all cases of glioblastoma multiforme overexpress the neurokinin type 1 receptor; the natural ligand is substance P, which was metabolically stabilized, labeled with 90Y and 213Bi, and injected into resection cavities or directly into tumors, which were critically located via a catheter system. The major advantage of this approach appeared to be the facilitated resectability of tumors, particularly in those patients who had been treated up front with the locoregional approach. It appears that neoadjuvant treatment before resection is a valid concept. Finally, another peptide family, the arginine-glycine-aspartate–based radiotracers, has made it to the clinic labeled with a variety of radioisotopes for monitoring the integrins αvβ3 overexpressed during tumor angiogenesis.

Clinical predictors and algorithm for the genetic diagnosis of pheochromocytoma patients.

Purpose: Six pheochromocytoma susceptibility genes causing distinct syndromes have been identified; approximately one of three of all pheochromocytoma patients carry a predisposing germline mutation. When four major genes (VHL, RET, SDHB, SDHD) are analyzed in a clinical laboratory, costs are ∼

Short‐term dietary restriction and fasting precondition against ischemia reperfusion injury in mice

3,400 per patient. The aim of the study is to systematically obtain a robust algorithm to identify who should be genetically tested, and to determine the order in which genes should be tested. Experimental Design: DNA from 989 apparently nonsyndromic patients were scanned for germline mutations in the genes VHL, RET, SDHB, SDHC, and SDHD. Clinical parameters were analyzed as potential predictors for finding mutations by multiple logistic regression, validated by bootstrapping. Cost reduction was calculated between prioritized gene testing compared with that for all genes. Results: Of 989 apparently nonsyndromic pheochromocytoma cases, 187 (19%) harbored germline mutations. Predictors for presence of mutation are age <45 years, multiple pheochromocytoma, extra-adrenal location, and previous head and neck paraganglioma. If we used the presence of any one predictor as indicative of proceeding with gene testing, then 342 (34.6%) patients would be excluded, and only 8 carriers (4.3%) would be missed. We were also able to statistically model the priority of genes to be tested given certain clinical features. E.g., for patients with prior head and neck paraganglioma, the priority would be SDHD>SDHB>RET>VHL. Using the clinical predictor algorithm to prioritize gene testing and order, a 44.7% cost reduction in diagnostic process can be achieved. Conclusions: Clinical parameters can predict for mutation carriers and help prioritize gene testing to reduce costs in nonsyndromic pheochromocytoma presentations. (Clin Cancer Res 2009;15(20):6378–85)

Evaluation of a 1,4,7,10-Tetraazacyclododecane-1,4,7,10-Tetraacetic Acid–Conjugated Bombesin-Based Radioantagonist for the Labeling with Single-Photon Emission Computed Tomography, Positron Emission Tomography, and Therapeutic Radionuclides

Dietary restriction (DR) extends lifespan and increases resistance to multiple forms of stress, including ischemia reperfusion injury to the brain and heart in rodents. While maximal effects on lifespan require long‐term restriction, the kinetics of onset of benefits against acute stress is not known. Here, we show that 2–4 weeks of 30% DR improved survival and kidney function following renal ischemia reperfusion injury in mice. Brief periods of water‐only fasting were similarly effective at protecting against ischemic damage. Significant protection occurred within 1 day, persisted for several days beyond the fasting period and extended to another organ, the liver. Protection by both short‐term DR and fasting correlated with improved insulin sensitivity, increased expression of markers of antioxidant defense and reduced expression of markers of inflammation and insulin/insulin‐like growth factor‐1 signaling. Unbiased transcriptional profiling of kidneys from mice subject to short‐term DR or fasting revealed a significant enrichment of signature genes of long‐term DR. These data demonstrate that brief periods of reduced food intake, including short‐term daily restriction and fasting, can increase resistance to ischemia reperfusion injury in rodents and suggest a rapid onset of benefits of DR in mammals.

Glucagon-Like Peptide-1 Versus Somatostatin Receptor Targeting Reveals 2 Distinct Forms of Malignant Insulinomas

Purpose: G protein–coupled receptor agonists are being used as radiolabeled vectors for in vivo localization and therapy of tumors. Recently, somatostatin-based antagonists were shown to be superior to agonists. Here, we compare the new [111In/68Ga]-labeled bombesin-based antagonist RM1 with the agonist [111In]-AMBA for targeting the gastrin-releasing peptide receptor (GRPR). Experimental Design: IC50, Kd values, and antagonist potency were determined using PC-3 and HEK-GRPR cells. Biodistribution and imaging studies were done in nude mice transplanted with the PC-3 tumor. The antagonist potency was assessed by evaluating the effects on calcium release and on receptor internalization monitored by immunofluorescence microscopy. Results: The IC50 value of [natIn]-RM1 was 14 ± 3.4 nmol/L. [nat/111In]-RM1 was found to bind to the GRPR with a Kd of 8.5 ± 2.7 nmol/L compared with a Kd of 0.6 ± 0.3 nmol/L of [111In]-AMBA. A higher maximum number of binding site value was observed for [111In]-RM1 (2.4 ± 0.2 nmol/L) compared with [111In]-AMBA (0.7 ± 0.1 nmol/L). [natLu]-AMBA is a potent agonist in the immunofluorescence-based internalization assay, whereas [natIn]-RM1 is inactive alone but efficiently antagonizes the bombesin effect. These data are confirmed by the calcium release assay. The pharmacokinetics showed a superiority of the radioantagonist with regard to the high tumor uptake (13.4 ± 0.8% IA/g versus 3.69 ± 0.75% IA/g at 4 hours after injection. as well as to all tumor-to-normal tissue ratios. Conclusion: Despite their relatively low GRPR affinity, the antagonists [111In/68Ga]-RM1 showed superior targeting properties compared with [111In]-AMBA. As found for somatostatin receptor–targeting radiopeptides, GRP-based radioantagonists seem to be superior to radioagonists for in vivo imaging and potentially also for targeted radiotherapy of GRPR-positive tumors. (Clin Cancer Res 2009;15(16):5240–9)

EANM procedure guideline for the treatment of liver cancer and liver metastases with intra-arterial radioactive compounds

Glucagon-like peptide-1 (GLP-1) receptor imaging is superior to somatostatin receptor subtype 2 (sst2) imaging in localizing benign insulinomas. Here, the role of GLP-1 and sst2 receptor imaging in the management of malignant insulinoma patients was investigated. Methods: Eleven patients with malignant insulinoma were prospectively included. 111In-[Lys40(Ahx-diethylenetriaminepentaacetic acid [DTPA])NH2]-exendin-4 SPECT/CT, 68Ga- DOTATATE PET/CT, and in vitro receptor autoradiography were performed to assess the receptor status and to evaluate the detection rate. Results: GLP-1 receptor targeting was positive in 4 of 11 patients, and sst2 receptor expression was positive in 8 of 11. In only 1 patient were both receptors expressed. In 1 patient, GLP-1 receptor imaging was the only method that successfully localized the primary tumor in the pancreas. In 3 patients with sst2-expressing tumors, DOTATATE radiotherapy was effectively applied. Conclusion: As opposed to benign insulinomas, malignant insulinomas often lack GLP-1 receptors. Conversely, malignant insulinomas often express sst2, which can be targeted therapeutically.

Glucagon-like peptide-1 receptor imaging for the localisation of insulinomas: a prospective multicentre imaging study

Primary liver cancers (i.e. hepatocellular carcinoma or cholangiocarcinoma) are worldwide some of the most frequent cancers, with rapidly fatal liver failure in a large majority of patients. Curative therapy consists of surgery (i.e. resection or liver transplantation), but only 10–20% of patients are candidates for this. In other patients, a variety of palliative treatments can be given, such as chemoembolization, radiofrequency ablation or recently introduced tyrosine kinase inhibitors, e.g. sorafenib. Colorectal cancer is the second most lethal cancer in Europe and liver metastases are prevalent either at diagnosis or in follow-up. These patients are usually treated by a sequence of surgery, chemotherapy and antibody therapy [Okuda et al. (Cancer 56:918–928, 1985); Schafer and Sorrell (Lancet 353:1253–1257, 1999); Leong et al. (Arnold, London, 1999)]. Radioembolization is an innovative therapeutic approach defined as the injection of micron-sized embolic particles loaded with a radioisotope by use of percutaneous intra-arterial techniques. Advantages of the use of these intra-arterial radioactive compounds are the ability to deliver high doses of radiation to small target volumes, the relatively low toxicity profile, the possibility to treat the whole liver including microscopic disease and the feasibility of combination with other therapy modalities. Disadvantages are mainly due to radioprotection constraints mainly for 131I-labelled agents, logistics and the possibility of inadvertent delivery or shunting [Novell et al. (Br J Surg 78:901–906, 1991)]. The Therapy, Oncology and Dosimetry Committees have worked together in order to revise the European Association of Nuclear Medicine (EANM) guidelines on the use of the radiopharmaceutical 131I-Lipiodol (Lipiocis®, IBA, Brussels, Belgium) and include the newer medical devices with 90Y-microspheres. 90Y is either bound to resin (SIR-Spheres®, Sirtex Medical, Lane Cove, Australia) or embedded in a glass matrix (TheraSphere®, MDS Nordion, Kanata, ON, Canada). Since 90Y-microspheres are not metabolized, they are not registered as unsealed sources. However, the microspheres are delivered in aqueous solution: radioactive contamination is a concern and microspheres should be handled, like other radiopharmaceuticals, as open sources. The purpose of this guideline is to assist the nuclear medicine physician in treating and managing patients undergoing such treatment.

Bombesin Antagonist–Based Radioligands for Translational Nuclear Imaging of Gastrin-Releasing Peptide Receptor–Positive Tumors

BACKGROUND Small benign insulinomas are hard to localise, leading to difficulties in planning of surgical interventions. We aimed to prospectively assess the insulinoma detection rate of single-photon emission CT in combination with CT (SPECT/CT) with a glucagon-like peptide-1 receptor avid radiotracer, and compare detection rates with conventional CT/MRI techniques. METHODS In our prospective imaging study, we enrolled adults aged 25-81 years at centres in Germany, Switzerland, and the UK. Eligible patients had proven clinical and biochemical endogenous hyperinsulinaemic hypoglycaemia and no evidence for metastatic disease on conventional imaging. CT/MRI imaging was done at referring centres according to standard protocols. At three tertiary nuclear medicine centres, we used whole body planar images and SPECT/CT of the abdomen up to 168 h after injection of (111)In-[Lys40(Ahx-DTPA-(111)In)NH2]-exendin-4 ((111)In-DTPA-exendin-4) to identify insulinomas. Consenting patients underwent surgery and imaging findings were confirmed histologically. FINDINGS Between Oct 1, 2008, and Dec 31, 2011, we recruited 30 patients. All patients underwent (111)In-DTPA-exendin-4 imaging, 25 patients underwent surgery (with histological analysis), and 27 patients were assessed with CT/MRI. (111)In-DTPA-exendin-4 SPECT/CT correctly detected 19 insulinomas and four additional positive lesions (two islet-cell hyperplasia and two uncharacterised lesions) resulting in a positive predictive value of 83% (95% CI 62-94). One true negative (islet-cell hyperplasia) and one false negative (malignant insulinoma) result was identified in separate patients by (111)In-DTPA-exendin-4 SPECT/CT. Seven patients (23%) were referred to surgery on the basis of (111)In-DTPA-exendin-4 imaging alone. For 23 assessable patients, (111)In-DTPA-exendin-4 SPECT/CT had a higher sensitivity (95% [95% CI 74-100]) than did CT/MRI (47% [27-68]; p=0.011). INTERPRETATION (111)In-DTPA-exendin-4 SPECT/CT could provide a good second-line imaging strategy for patients with negative results on initial imaging with CT/MRI. FUNDING Oncosuisse, the Swiss National Science Foundation, and UK Department of Health.

SPECT Study of Folate Receptor-Positive Malignant and Normal Tissues in Mice Using a Novel 99mTc-Radiofolate

Bombesin receptors are overexpressed on a variety of human tumors. In particular, the gastrin-releasing peptide receptor (GRPr) has been identified on prostate and breast cancers and on gastrointestinal stromal tumors. The current study aims at developing clinically translatable bombesin antagonist–based radioligands for SPECT and PET of GRPr-positive tumors. Methods: A potent bombesin antagonist (PEG4-d-Phe-Gln-Trp-Ala-Val-Gly-His-Sta-Leu-NH2 [AR]) was synthesized; conjugated to the chelators DOTA, 6-carboxy-1,4,7,11-tetraazaundecane (N4), 1,4,7-triazacyclononane, 1-glutaric acid-4,7 acetic acid (NODAGA), and 4,11-bis(carboxymethyl)-1,4,8,11-tetraazabicyclo[6.6.2]hexadecane (CB-TE2A); and radiolabeled with 111In, 99mTc, 68Ga, and 64Cu, respectively. The radioconjugates were evaluated in vitro and in vivo in PC-3 tumor–bearing nude mice. Antagonist potency was determined by Ca2+-flux measurements and immunofluorescence. Results: All the conjugates showed high binding affinity to GRPr (inhibitory concentration of 50% [IC50], 2.5–25 nmol/L). The immunofluorescence and Ca2+-flux assays confirmed the antagonist properties of the conjugates. Biodistribution revealed high and specific uptake in PC-3 tumor and in GRPr-positive tissues. Tumor uptake of 64Cu-CB-TE2A-AR (31.02 ± 3.35 percentage injected activity per gram [%IA/g]) was higher than 99mTc-N4-AR (24.98 ± 5.22 %IA/g), 111In-DOTA-AR (10.56 ± 0.70 %IA/g), and 68Ga-NODAGA-AR (7.11 ± 3.26 %IA/g) at 1 h after injection. Biodistribution at later time points showed high tumor-to-background ratios because of the fast washout of the radioligand from normal organs, compared with tumor. High tumor-to-background ratios were further illustrated by PET and SPECT images of PC-3 tumor–bearing nude mice acquired at 12 h after injection showing high tumor uptake, clear background, and negligible or no radioactivity in the abdomen. Conclusion: The chelators do influence the affinity, antagonistic potency, and pharmacokinetics of the conjugates. The promising preclinical results warrant clinical translation of these probes for SPECT and PET.