Helmut R. Mäcke

Affinity profiles for human somatostatin receptor subtypes SST1-SST5 of somatostatin radiotracers selected for scintigraphic and radiotherapeutic use.

Abstract.In vivo somatostatin receptor scintigraphy using Octreoscan is a valuable method for the visualisation of human endocrine tumours and their metastases. Recently, several new, alternative somatostatin radioligands have been synthesised for diagnostic and radiotherapeutic use in vivo. Since human tumours are known to express various somatostatin receptor subtypes, it is mandatory to assess the receptor subtype affinity profile of such somatostatin radiotracers. Using cell lines transfected with somatostatin receptor subtypes sst1, sst2, sst3, sst4 and sst5, we have evaluated the in vitro binding characteristics of labelled (indium, yttrium, gallium) and unlabelled DOTA-[Tyr3]-octreotide, DOTA-octreotide, DOTA-lanreotide, DOTA-vapreotide, DTPA-[Tyr3]-octreotate and DOTA-[Tyr3]-octreotate. Small structural modifications, chelator substitution or metal replacement were shown to considerably affect the binding affinity. A marked improvement of sst2 affinity was found for Ga-DOTA-[Tyr3]-octreotide (IC50 2.5 nM) compared with the Y-labelled compound and Octreoscan. An excellent binding affinity for sst2 in the same range was also found for In-DTPA-[Tyr3]-octreotate (IC50 1.3 nM) and for Y-DOTA-[Tyr3]-octreotate (IC50 1.6 nM). Remarkably, Ga-DOTA-[Tyr3]-octreotate bound at sst2 with a considerably higher affinity (IC50 0.2 nM). An up to 30-fold improvement in sst3 affinity was observed for unlabelled or Y-labelled DOTA-octreotide compared with their Tyr3-containing analogue, suggesting that replacement of Tyr3 by Phe is crucial for high sst3 affinity. Substitution in the octreotide molecule of the DTPA by DOTA improved the sst3 binding affinity 14-fold. Whereas Y-DOTA-lanreotide had only low affinity for sst3 and sst4, it had the highest affinity for sst5 among the tested compounds (IC50 16 nM). Increased binding affinity for sst3 and sst5 was observed for DOTA-[Tyr3]-octreotide, DOTA-lanreotide and DOTA-vapreotide when they were labelled with yttrium. These marked changes in subtype affinity profiles are due not only to the different chemical structures but also to the different charges and hydrophilicity of these compounds. Interestingly, even the coordination geometry of the radiometal complex remote from the pharmacophoric amino acids has a significant influence on affinity profiles as shown with Y-DOTA versus Ga-DOTA in either [Tyr3]-octreotide or [Tyr3]-octreotate. Such changes in sst affinity profiles must be identified in newly designed radiotracers used for somatostatin receptor scintigraphy in order to correctly interpret in vivo scintigraphic data. These observations may represent basic principles relevant to the development of other peptide radioligands.

Response, Survival, and Long-Term Toxicity After Therapy With the Radiolabeled Somatostatin Analogue [90Y-DOTA]-TOC in Metastasized Neuroendocrine Cancers

PURPOSE To investigate response, survival, and safety profile of the somatostatin-based radiopeptide (90)yttrium-labeled tetraazacyclododecane-tetraacetic acid modified Tyr-octreotide ([(90)Y-DOTA]-TOC) in neuroendocrine cancers. PATIENTS AND METHODS In a clinical phase II single-center open-label trial, patients with neuroendocrine cancers were treated with repeated cycles of [(90)Y-DOTA]-TOC. Each cycle consisted of a single intravenous injection of 3.7GBq/m(2) body-surface [(90)Y-DOTA]-TOC. Additional cycles were withheld in case of tumor progression and/or permanent toxicity. RESULTS Overall, 1,109 patients received 2,472 cycles of [(90)Y-DOTA]-TOC (median, two; range, one to 10 cycles per patient). Of the 1,109 patients, 378 (34.1%) experienced morphologic response; 172 (15.5%), biochemical response; and 329 (29.7%), clinical response. During a median follow-up of 23 months, 491 patients (44.3%) died. Longer survival was correlated with each: morphologic (hazard ratio [HR], 0.46; 95% CI, 0.38 to 0.56; median survival, 44.7 v 18.3 months; P < .001), biochemical (HR, 0.75; 95% CI, 0.59 to 0.96; 35.3 v 25.7 months; P = .023), and clinical response (HR, 0.68; 95% CI, 0.56 to 0.82; 36.8 v 23.5 months; P < .001). Overall, 142 patients (12.8%) developed grade 3 to 4 transient hematologic toxicities, and 103 patients (9.2%) experienced grade 4 to 5 permanent renal toxicity. Multivariable regression revealed that tumoral uptake in the initial imaging study was predictive for overall survival (HR, 0.45; 95% CI, 0.29 to 0.69; P < .001), whereas the initial kidney uptake was predictive for severe renal toxicity (HR, 1.59; 95% CI, 1.17 to 2.17; P = .003). CONCLUSION This study documents the long-term outcome of [(90)Y-DOTA]-TOC treatment in a large cohort. Response to [(90)Y-DOTA]-TOC is associated with longer survival. Somatostatin receptor imaging is predictive for both survival after [(90)Y-DOTA]-TOC treatment and occurrence of renal toxicity.

Evaluation of Positron Emission Tomography Imaging Using [68Ga]-DOTA-D Phe1-Tyr3-Octreotide in Comparison to [111In]-DTPAOC SPECT. First Results in Patients with Neuroendocrine Tumors

PURPOSE [111In]-DTPAOC (Octreoscan(R)) has been shown to be very useful in the detection of somatostatin receptor (SSTR) positive tumors and their metastases using either conventional scintigraphy or single photon emission computed tomography (SPECT). The main drawback of this method is the limited spatial resolution and a somewhat low receptor affinity of the radiopeptide. Due to the increased spatial resolution and the ability of quantification, an agent for positron emission tomography (PET) imaging of SSTR is desirable. This communication shows our initial experience using [68Ga]-DOTA-D-Phe(1)-Tyr(3)-Octreotide (DOTATOC) in comparison to [111In]-DTPAOC-SPECT in patients with neuroendocrine tumors. PROCEDURES Four patients, two male and two female (46-55 years old) have been examined by [111In]-DTPAOC scintigraphy and within one month by [68Ga]-DOTATOC-PET. All of them suffered from neuroendocrine tumors and/or their metastases. DOTATOC has been labeled using the positron-emitting generator-nuclide 68Ga (t(1/2) 68 minutes). In two patients with previously known localization of tumor, dynamic PET scans after intravenous bolus-injection of 181+/-17 MBq [68Ga]-DOTATOC until 120 minutes post-injection were acquired. In all patients, the static PET-scans have been acquired after 45 or 60 minutes post-injection (SUV1) and 140 minutes post-injection (SUV2). RESULTS Similar to [111In]-DTPAOC, [68Ga]-DOTATOC showed the highest uptake in the spleen, followed by the kidneys and the liver. A clear delineation of the pituitary gland could only be achieved by PET. The highest SUVs were found at a plateau between 45 and 90 minutes with a maximum 60 minutes post-injection. Due to the fast tracer accumulation in the tumor and the rapid clearance of the compound, resulting in high tumor to background ratios even 40 minutes after injection, the short half life of 68Ga is reasonable. In two patients more findings have been revealed by [68Ga]-DOTATOC-PET as compared to the [111In]-DTPAOC-SPECT. In comparison to the [111In]-DTPAOC-SPECT [68Ga]-DOTATOC-PET seems to be superior especially concerning small findings with low tracer uptake. Both [111In]-DTPAOC-SPECT and [68Ga]-DOTATOC-PET were less sensitive in the detection of liver metastases of neuroendocrine tumors compared to computerized tomography CT because they showed a lower uptake than the surrounding liver tissue. CONCLUSIONS According to our initial experiences in a limited number of patients, [68Ga]-DOTATOC is a promising PET tracer for imaging neuroendocrine tumors and their metastases. In comparison to the [111In]-DTPAOC-scan it seems to be superior especially in detecting small tumors or tumors bearing only a low density of SSTRs. It offers excellent imaging properties and very high tumor to background ratios. Further evaluation of [68Ga]-DOTATOC in a larger number of patients is certainly justified.

Radiolabeled somatostatin receptor antagonists are preferable to agonists for in vivo peptide receptor targeting of tumors

Targeting neuroendocrine tumors expressing somatostatin receptor subtypes (sst) with radiolabeled somatostatin agonists is an established diagnostic and therapeutic approach in oncology. While agonists readily internalize into tumor cells, permitting accumulation of radioactivity, radiolabeled antagonists do not, and they have not been considered for tumor targeting. The macrocyclic chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) was coupled to two potent somatostatin receptor-selective peptide antagonists [NH2-CO-c(DCys-Phe-Tyr-DAgl8(Me,2-naphthoyl)-Lys-Thr-Phe-Cys)-OH (sst3-ODN-8) and a sst2-selective antagonist (sst2-ANT)], for labeling with 111/natIn. 111/natIn-DOTA-sst3-ODN-8 and 111/natIn-DOTA–[4-NO2-Phe-c(DCys-Tyr-DTrp-Lys-Thr-Cys)-DTyr-NH2] (111/natIn-DOTA-sst2-ANT) showed high sst3- and sst2-binding affinity, respectively. They did not trigger sst3 or sst2 internalization but prevented agonist-stimulated internalization. 111In-DOTA-sst3-ODN-8 and 111In-DOTA-sst2-ANT were injected intravenously into mice bearing sst3- and sst2-expressing tumors, and their biodistribution was monitored. In the sst3-expressing tumors, strong accumulation of 111In-DOTA-sst3-ODN-8 was observed, peaking at 1 h with 60% injected radioactivity per gram of tissue and remaining at a high level for >72 h. Excess of sst3-ODN-8 blocked uptake. As a control, the potent agonist 111In-DOTA–[1-Nal3]-octreotide, with strong sst3-binding and internalization properties showed a much lower and shorter-lasting uptake in sst3-expressing tumors. Similarly, 111In-DOTA-sst2-ANT was injected into mice bearing sst2-expressing tumors. Tumor uptake was considerably higher than with the highly potent sst2-selective agonist 111In-diethylenetriaminepentaacetic acid–[Tyr3,Thr8]-octreotide (111In-DTPA-TATE). Scatchard plots showed that antagonists labeled many more sites than agonists. Somatostatin antagonist radiotracers therefore are preferable over agonists for the in vivo targeting of sst3- or sst2-expressing tumors. Antagonist radioligands for other peptide receptors need to be evaluated in nuclear oncology as a result of this paradigm shift.

Yttrium-90 and indium-111 labelling, receptor binding and biodistribution of [DOTA0,d-Phe1,Tyr3]octreotide, a promising somatostatin analogue for radionuclide therapy

In vitro octreotide receptor binding of [111In-DOTA0,d-Phe1,Tyr3]octreotide (111In-DOTATOC) and the in vivo metabolism of90Y or111In-labelled DOTATOC were investigated in rats in comparison with [111In-DTPA0]octreotide [111In-DTPAOC).111In-DOTATOC was found to have an affinity similar to octreotide itself for the octreotide receptor in rat cerebral cortex microsomes. Twenty-four hours after injection of90Y or111In-labelled DOTATOC, uptake of radioactivity in the octreotide receptor-expressing tissues pancreas, pituitary, adrenals and tumour was a factor of 2–6 that after injection of111In-DTPAOC. Uptake of labelled DOTATOC in pituitary, pancreas, adrenals and tumour was almost completely blocked by pretreatment with 0.5 mg unlabelled octreotide, indicating specific binding to the octreotide receptors. These findings strongly indicate that90Y-DOTATOC is a promising radiopharmaceutical for radiotherapy and that111In-DOTATOC is of potential value for diagnosis of patients with octreotide receptor-positive lesions, such as most neuroendocrine tumours.

Glucagon-like peptide 1-receptor scans to localize occult insulinomas

The precise localization of some insulinomas with the use of conventional imaging techniques is a challenging clinical problem. These findings indicate that GLP-1–receptor scanning may offer a new ...

Exendin-4–Based Radiopharmaceuticals for Glucagonlike Peptide-1 Receptor PET/CT and SPECT/CT

Strong overexpression of glucagonlike peptide-1 (GLP-1) receptors in human insulinoma provides an attractive target for imaging. The first clinical trials demonstrated that GLP-1 receptor SPECT/CT using [Lys40(Ahx [6-aminohexanoic acid]-DOTA-111In)NH2]-exendin-4 can localize hardly detectable insulinomas. However, [Lys40(Ahx-DOTA-111In)NH2]-exendin-4 imaging has drawbacks related to the use of 111In in that it is costly and carries a relatively high radiation burden for the patient. The aim of this study was the preclinical evaluation of [Lys40(Ahx-DOTA-68Ga)NH2]-exendin-4 for PET/CT and [Lys40(Ahx-hydrazinonicotinamide [HYNIC]-99mTc)NH2]-exendin-4 for SPECT/CT. Methods: Internalization, biodistribution, dosimetry, and imaging studies were performed in the Rip1Tag2 mouse model of pancreatic β-cell carcinogenesis and compared with our gold standard [Lys40(Ahx-DOTA-111In)NH2]-exendin-4. Poly-glutamic acid and Gelofusine, a gelatin-based plasma expander, were used for renal uptake reduction studies. Results: The tumor uptake of [Lys40(Ahx-DOTA-68Ga)NH2]-exendin-4 was 205 ± 59 percentage injected activity per gram of tissue at 4 h. Other GLP-1 receptor–positive organs showed more than 4.8 times lower radioactivity uptake. [Lys40(Ahx-HYNIC-99mTc/ethylenediaminediacetic acid [EDDA])NH2]-exendin-4, compared with its 111In- and 68Ga-labeled sister compounds, showed significantly less tumor and organ uptake. The significantly lower tumor and organ uptake of [Lys40(Ahx-HYNIC-99mTc/EDDA)NH2]-exendin-4 did not result in inferior tumor-to-organ ratios or reduced image quality. All radiopeptides tested showed a high tumor-to-background ratio, resulting in the visualization of small tumors (maximum diameter between 1.0 and 3.2 mm) by SPECT and PET. The only exception was the kidneys, which also showed high uptake. This uptake could be reduced by 49%−78% using poly-glutamic acid, Gelofusine, or a combination of the 2. The estimated effective radiation dose was 3.7 μSv/MBq for [Lys40(Ahx-HYNIC-99mTc/EDDA)NH2]-exendin-4, which was 8 times less than that for [Lys40(Ahx-DOTA-68Ga)NH2]-exendin-4 and 43 times less than that for [Lys40(Ahx-DOTA-111In)NH2]-exendin-4. Conclusion: These promising pharmacokinetic and imaging data show that [Lys40(Ahx-DOTA-68Ga)NH2]-exendin-4 and [Lys40(Ahx-HYNIC-99mTc/EDDA)NH2]-exendin-4 are suitable candidates for clinical GLP-1 receptor imaging studies.

Internalization of radiolabelled [DTPA0]octreotide and [DOTA0,Tyr3]octreotide: peptides for somatostatin receptor-targeted scintigraphy and radionuclide therapy.

We compared the internalization of [90Y-DOTA0,Tyr3]octreotide and [111In-DOTA0,Tyr3]octreotide with that of [125I-Tyr3]octreotide and [111In-DTPA0]octreotide in the subtype 2 somatostatin receptor (sst2)-positive rat pancreatic tumour cell lines CA20948 and AR42J and in the somatostatin receptor-negative human anaplastic thyroid tumour cell line ARO. We demonstrated that [111In-DTPA0]octreotide, [90Y-DOTA0,Tyr3]octreotide and [111In-DOTA0,Tyr3]octreotide are internalized by a receptor-specific, time- and temperature-dependent process. The amount of [90Y-DOTA0,Tyr3]octreotide internalized was higher than that of [111In-DOTA0,Tyr3]octreotide and [111In-DTPA0]octreotide.

Cohort Study of Somatostatin-Based Radiopeptide Therapy With ( 90 Y-DOTA)-TOC Versus ( 90 Y-DOTA)-TOC Plus ( 177 Lu-DOTA)-TOC in Neuroendocrine Cancers

PURPOSE Radiopeptide therapy is commonly performed with a single radioisotope. We aimed to compare the effectiveness of somatostatin-based radiopeptide therapy with a single versus a combination of radioisotopes. PATIENTS AND METHODS In a cohort study, patients with metastasized neuroendocrine cancer were treated with repeated cycles of (90)yttrium-labeled tetraazacyclododecane-tetraacetic acid modified Tyr-octreotide ([(90)Y-DOTA]-TOC) or with cycles alternating between [(90)Y-DOTA]-TOC and (177)lutetium-labeled DOTA-TOC ([(177)Lu-DOTA]-TOC) until tumor progression or permanent toxicity. Multivariable Cox regression and competing risk regression were used to study predictors of survival and renal toxicity in patients completing three or more treatment cycles. RESULTS A total of 486 patients completed three or more treatment cycles; 237 patients received [(90)Y-DOTA]-TOC and 249 patients received [(90)Y-DOTA]-TOC + [(177)Lu-DOTA]-TOC. Patients receiving [(90)Y-DOTA]-TOC + [(177)Lu-DOTA]-TOC had a significantly longer survival than patients receiving [(90)Y-DOTA]-TOC alone (5.51 v 3.96 years; hazard ratio, 0.64; 95% CI, 0.47 to 0.88; P = .006). The rates of severe hematologic toxicities (6.3% v 4.4%; P = .25) and severe renal toxicity (8.9% v 11.2%; P = .47) were comparable in both groups. CONCLUSION [(90)Y-DOTA]-TOC + [(177)Lu-DOTA]-TOC was associated with improved overall survival compared with [(90)Y-DOTA]-TOC alone in patients completing three or more cycles of treatment. Contrary to the current practice in radiopeptide therapy, our results suggest an advantage of using a combination of radioisotopes.

Pre-clinical comparison of [DTPA0] octreotide, [DTPA0,Tyr3] octreotide and [DOTA0,Tyr3] octreotide as carriers for somatostatin receptor-targeted scintigraphy and radionuclide therapy

We have evaluated the potential usefulness of radiolabelled [DTPA0,Tyr3]octreotide and [DOTA0,Tyr3]octreotide as radiopharmaceuticals for somatostatin receptor–targeted scintigraphy and radiotherapy. In vitro somatostatin receptor binding and in vivo metabolism in rats of the compounds were investigated in comparison with [111In‐DTPA0] octreotide. Comparing different peptide–chelator constructs, [DTPA0,Tyr3]octreotide and [DOTA0, Tyr3]octreotide were found to have a higher affinity than [DTPA0]octreotide for subtype 2 somatostatin receptors (sst2) in mouse AtT20 pituitary tumour cell membranes (all IC50 values obtained were in the low nanomolar range). In vivo studies in CA20948 tumor‐bearing Lewis rats revealed a significantly higher uptake of both 111In‐labelled [DOTA0,Tyr3]octreotide and [DTPA0,Tyr3]octreotide in sst2‐expressing tissues than after injection of [111In‐DTPA0]octreotide, showing that substitution of Tyr for Phe at position 3 in octreotide results in an increased affinity for its receptor and in a higher target tissue uptake. Uptake of 111In‐labelled [DTPA0]octreotide, [DTPA0,Tyr3]octreotide and [DOTA0,Tyr3]octreotide in pituitary, pancreas, adrenals and tumour was decreased to less than 7% of control by pre‐treatment with 0.5 mg unlabelled octreotide/rat, indicating specific binding to sst2. Comparing different radionuclides, [90Y‐DOTA0,Tyr3]octreotide had the highest uptake in sst2‐positive organs, followed by the [111In‐DOTA0,Tyr3]octreotide, whereas [DOTA0, 125I‐Try3]octreotide uptake was low compared to that of the other radiopharmaceuticals, when measured 24 hr after injection. Renal uptake of 111In‐labelled [DTPA0]octreotide, [DTPA0, Tyr3]octreotide and [DOTA0,Tyr3]octreotide was reduced over 50% by an i.v. injection of 400 mg/kg d‐lysine, whereas radioactivity in blood, pancreas and adrenals was not affected. Int. J. Cancer 75:406–411, 1998.