Somatostatin Receptor Antagonist Research Articles

Impressive Response to TANDEM Peptide Receptor Radionuclide Therapy with 177Lu/225AcDOTA-LM3 Somatostatin Receptor Antagonist in a Patient with Therapy-Refractory, Rapidly Progressive Neuroendocrine Neoplasm of the Pancreas.

The present report describes the history of a 58-year-old woman with a rapidly progressing neuroendocrine pancreatic tumor (initially G2) presenting with extensive liver, bone, and lymph node metastases. Previous treatments included chemotherapy, hemithyroidectomy for right lobe metastasis, Peptide Receptor Radionuclide Therapy (PRRT) with [177Lu]Lu-DOTATATE, Lanreotide, Everolimus, and liver embolization. Due to severe disease progression, after a liver biopsy revealing tumor grade G3, PRRT with the somatostatin receptor antagonist LM3 was initiated. [68Ga]GaDOTA-LM3 PET/CT showed intense tracer uptake in the liver, pancreatic tumor, lymph nodes, and bone metastases. Three TANDEM-PRRT cycles using [177Lu]LuDOTA-LM3 and [225Ac]AcDOTA-LM3, administered concurrently, resulted in significant improvement, notably in liver metastases, hepatomegaly reduction, the complete regression of bone and lymph node metastases, and primary tumor improvement. Partial remission was confirmed by positron emission tomography/computed tomography, chest-abdomen-pelvis contrast-enhanced computed tomography, and magnetic resonance of the abdomen, with marked clinical improvement in pain, energy levels, and quality of life, enabling full resumption of physical activity.

Dosimetry and pharmacokinetics of [177Lu]Lu-satoreotide tetraxetan in patients with progressive neuroendocrine tumours.

To evaluate the dosimetry and pharmacokinetics of the novel radiolabelled somatostatin receptor antagonist [177Lu]Lu-satoreotide tetraxetan in patients with advanced neuroendocrine tumours (NETs). This study was part of a phase I/II trial of [177Lu]Lu-satoreotide tetraxetan, administered at a median cumulative activity of 13.0 GBq over three planned cycles (median activity/cycle: 4.5 GBq), in 40 patients with progressive NETs. Organ absorbed doses were monitored at each cycle using patient-specific dosimetry; the cumulative absorbed-dose limits were set at 23.0Gy for the kidneys and 1.5Gy for bone marrow. Absorbed dose coefficients (ADCs) were calculated using both patient-specific and model-based dosimetry for some patients. In all evaluated organs, maximum [177Lu]Lu-satoreotide tetraxetan uptake was observed at the first imaging timepoint (4h after injection), followed by an exponential decrease. Kidneys were the main route of elimination, with a cumulative excretion of 57-66% within 48h following the first treatment cycle. At the first treatment cycle, [177Lu]Lu-satoreotide tetraxetan showed a median terminal blood half-life of 127h and median ADCs of [177Lu]Lu-satoreotide tetraxetan were 5.0Gy/GBq in tumours, 0.1Gy/GBq in the bone marrow, 0.9Gy/GBq in kidneys, 0.2Gy/GBq in the liver and 0.8Gy/GBq in the spleen. Using image-based dosimetry, the bone marrow and kidneys received median cumulative absorbed doses of 1.1 and 10.8Gy, respectively, after three cycles. [177Lu]Lu-satoreotide tetraxetan showed a favourable dosimetry profile, with high and prolonged tumour uptake, supporting its acceptable safety profile and promising efficacy. NCT02592707. Registered October 30, 2015.

Gallium-68 labeled somatostatin receptor antagonist PET/CT in over 500 patients with neuroendocrine neoplasms: experience from a single center in China.

Somatostatin receptor antagonists have shown promising performance for imaging neuroendocrine neoplasms. However, there is a lack of studies exploring the diagnostic performance of SSTR antagonists or comparing them with agonists in a large cohort of patients with NENs. This study aimed to retrospectively review all SSTR antagonist PET/CT scans conducted at Peking Union Medical College Hospital since November 2018 in patients with confirmed or suspected NENs. Four types of SSTR antagonists were utilized, including [68Ga]Ga-NODAGA-LM3, [68Ga]Ga-DOTA-LM3, [68Ga]Ga-NODAGA-JR11, and [68Ga]Ga-DOTA-JR11. The reference standard was based on a combination of histopathology, clinical evaluation, imaging results, and follow-up. Patient-based sensitivity, specificity, and accuracy were evaluated. The SUVmax and tumor-to-liver ratio (TLR) of the hottest lesions was recorded and compared between antagonists and [68Ga]Ga-DOTATATE. A total of 622 antagonist scans from 549 patients were included in the analysis. The patient-level sensitivity, specificity, and accuracy of antagonist imaging (all tracers combined) were 91.0% (443/487), 91.9% (57/62), and 91.1% (500/549), respectively. In 181 patients with a comparative [68Ga]Ga-DOTATATE PET/CT scan, the patient-level sensitivity, specificity, and accuracy were 87.5% (147/168), 76.9% (10/13), and 86.7% (157/181), respectively. For the hottest lesions, SSTR antagonists all tracers combined demonstrated an overall comparable SUVmax to [68Ga]Ga-DOTATATE (40.1 ± 32.5 vs. 39.4 ± 23.8, p = 0.772). While [68Ga]Ga-NODAGA-LM3 showed significantly higher uptake than [68Ga]Ga-DOTATATE (57.4 ± 38.5 vs. 40.0 ± 22.8, p<0.001), [68Ga]Ga-NODAGA-JR11 (39.7 ± 26.5 vs. 34.3 ± 23.9, p = 0.108) and [68Ga]Ga-DOTA-LM3 (38.9 ± 32.1 vs. 37.2 ± 22.1, p = 0.858) showed comparable uptake to [68Ga]Ga-DOTATATE, and [68Ga]Ga-DOTA-JR11 showed lower uptake (28.9 ± 26.1 vs. 44.0 ± 25.7, p = 0.001). All antagonists exhibited significantly higher TLR than [68Ga]Ga-DOTATATE (12.1 ± 10.8 vs. 5.2 ± 4.5, p<0.001). Gallium-68 labeled SSTR antagonists could serve as alternatives to SSTR agonists for imaging of NENs. Among various antagonists, [68Ga]Ga-NODAGA-LM3 seems to have the best imaging profile.

A phase I/II study of the safety and efficacy of [177Lu]Lu-satoreotide tetraxetan in advanced somatostatin receptor-positive neuroendocrine tumours

PurposeWe present the results of an open-label, phase I/II study evaluating the safety and efficacy of the novel somatostatin receptor (SSTR) antagonist [177Lu]Lu-satoreotide tetraxetan in 40 patients with previously treated, progressive neuroendocrine tumours (NETs), in which dosimetry was used to guide maximum administered activity.MethodsThis study was conducted in two parts. Part A consisted of 15 patients who completed three cycles of [177Lu]Lu-satoreotide tetraxetan at a fixed administered activity and peptide amount per cycle (4.5 GBq/300 µg). Part B, which included 25 patients who received one to five cycles of [177Lu]Lu-satoreotide tetraxetan, evaluated different administered activities (4.5 or 6.0 GBq/cycle) and peptide amounts (300, 700, or 1300 μg/cycle), limited to a cumulative absorbed radiation dose of 23 Gy to the kidneys and 1.5 Gy to the bone marrow.ResultsMedian cumulative administered activity of [177Lu]Lu-satoreotide tetraxetan was 13.0 GBq over three cycles (13.1 GBq in part A and 12.9 GBq in part B). Overall, 17 (42.5%) patients experienced grade ≥ 3 treatment‑related adverse events; the most common were lymphopenia, thrombocytopenia, and neutropenia. No grade 3/4 nephrotoxicity was observed. Two patients developed myeloid neoplasms considered treatment related by the investigator. Disease control rate for part A and part B was 94.7% (95% confidence interval [CI]: 82.3–99.4), and overall response rate was 21.1% (95% CI: 9.6–37.3).Conclusion[177Lu]Lu-satoreotide tetraxetan, administered at a median cumulative activity of 13.0 GBq over three cycles, has an acceptable safety profile with a promising clinical response in patients with progressive, SSTR-positive NETs. A 5-year long-term follow-up study is ongoing.Trial registrationClinicalTrials.gov, NCT02592707. Registered October 30, 2015.

Perspective of molecular imaging and peptide receptor radionuclide therapy in pancreatic neuroendocrine tumors: where do we stand?

The clinical use of nuclear medicine imaging and therapy in pancreatic neuroendocrine tumors has been greatly strengthened since the approval of 68Ga-DOTATATE and 177Lu-DOTATATE. However, many aspects are still under discussion. In this 2-part article, we aim to collect and discuss current evidence of molecular imaging and peptide receptor radionuclide therapy (PRRT) in pancreatic neuroendocrine tumor. In the first part, we will address some critical aspects of 68Ga-SSAs imaging, including diagnostic efficacy, recurrence detection and follow-up, patient selection for PRRT, and pitfalls in image interpretation. Besides, we will also briefly discuss the role of 18F-fluorodeoxyglucose positron emission tomography/computed tomography, special imaging strategy in regard to insulinoma, and the status of radiolabeled somatostatin receptor antagonist. In the second part, we aim to review the current evidence of PRRT in pancreatic neuroendocrine tumor, focusing on efficacy and safety in particular. We will also introduce the recent development of PRRT, including PRRT in high-grade neuroendocrine neoplasms, retreatment PRRT, upfront PRRT, PRRT in the setting of neoadjuvant therapy and conversion therapy, combination therapies with PRRT, PRRT with αradionuclides, and PRRT with antagonists.

Combination of terbium-161 with somatostatin receptor antagonists\u2014a potential paradigm shift for the treatment of neuroendocrine neoplasms

PurposeThe β¯-emitting terbium-161 also emits conversion and Auger electrons, which are believed to be effective in killing single cancer cells. Terbium-161 was applied with somatostatin receptor (SSTR) agonists that localize in the cytoplasm (DOTATOC) and cellular nucleus (DOTATOC-NLS) or with a SSTR antagonist that localizes at the cell membrane (DOTA-LM3). The aim was to identify the most favorable peptide/terbium-161 combination for the treatment of neuroendocrine neoplasms (NENs).MethodsThe capability of the 161Tb- and 177Lu-labeled somatostatin (SST) analogues to reduce viability and survival of SSTR-positive AR42J tumor cells was investigated in vitro. The radiopeptides’ tissue distribution profiles were assessed in tumor-bearing mice. The efficacy of terbium-161 compared to lutetium-177 was investigated in therapy studies in mice using DOTATOC or DOTA-LM3, respectively.ResultsIn vitro, [161Tb]Tb-DOTA-LM3 was 102-fold more potent than [177Lu]Lu-DOTA-LM3; however, 161Tb-labeled DOTATOC and DOTATOC-NLS were only 4- to fivefold more effective inhibiting tumor cell viability than their 177Lu-labeled counterparts. This result was confirmed in vivo and demonstrated that [161Tb]Tb-DOTA-LM3 was significantly more effective in delaying tumor growth than [177Lu]Lu-DOTA-LM3, thereby, prolonging survival of the mice. A therapeutic advantage of terbium-161 over lutetium-177 was also manifest when applied with DOTATOC. Since the nuclear localizing sequence (NLS) compromised the in vivo tissue distribution of DOTATOC-NLS, it was not used for therapy.ConclusionThe use of membrane-localizing DOTA-LM3 was beneficial and profited from the short-ranged electrons emitted by terbium-161. Based on these preclinical data, [161Tb]Tb-DOTA-LM3 may outperform the clinically employed [177Lu]Lu-DOTATOC for the treatment of patients with NENs.

A novel read methodology to evaluate the optimal dose of 68Ga-satoreotide trizoxetan as a PET imaging agent in patients with gastroenteropancreatic neuroendocrine tumours: a phase II clinical trial

Background68Ga-satoreotide trizoxetan is a novel somatostatin receptor antagonist exhibiting higher tumour-to-background ratios and sensitivity compared to 68Ga-DOTATOC. This randomised, 2 × 3 factorial, phase II study aimed to confirm the optimal peptide mass and radioactivity ranges for 68Ga-satoreotide trizoxetan, using binary visual reading. To that end, 24 patients with metastatic gastroenteropancreatic neuroendocrine tumours received 5–20 µg of 68Ga-satoreotide trizoxetan on day 1 of the study and 30–45 µg on day 16–22, with one of three gallium-68 radioactivity ranges (40–80, 100–140, or 160–200 MBq) per visit. Two 68Ga-satoreotide trizoxetan PET/CT scans were acquired from each patient post-injection, and were scored by experienced independent blinded readers using a binary system (0 for non-optimal image quality and 1 for optimal image quality). For each patient pair of 68Ga-satoreotide trizoxetan scans, one or both images could score 1.ResultsTotal image quality score for 68Ga-satoreotide trizoxetan PET scans was lower in the 40–80 MBq radioactivity range (56.3%) compared to 100–140 MBq (90.6%) and 160–200 MBq (81.3%). Both qualitative and semi-quantitative analysis showed that peptide mass (5–20 or 30–45 µg) did not influence 68Ga-satoreotide trizoxetan imaging. There was only one reading where readers diverged on scoring; one reader preferred one image because of higher lesion conspicuity, and the other reader preferred the alternative image because of the ability to identify more lesions.ConclusionsBinary visual reading, which was associated with a low inter-reader variability, has further supported that the optimal administered radioactivity of 68Ga-satoreotide trizoxetan was 100–200 MBq with a peptide mass up to 50 µg.Trial registration ClinicalTrials.gov, NCT03220217. Registered 18 July 2017, https://clinicaltrials.gov/ct2/show/NCT03220217

First-in-Humans Study of the SSTR Antagonist 177Lu-DOTA-LM3 for Peptide Receptor Radionuclide Therapy in Patients with Metastatic Neuroendocrine Neoplasms: Dosimetry, Safety, and Efficacy.

The objective of this study was to assess the safety, dosimetry, and efficacy of the 177Lu-labeled somatostatin receptor (SSTR) antagonist DOTA-p-Cl-Phe-cyclo(d-Cys-Tyr-d-4-amino-Phe(carbamoyl)-Lys-Thr-Cys)d-Tyr-NH2 (177Lu-DOTA-LM3) in patients with metastatic neuroendocrine neoplasms (NENs). Methods: Fifty-one patients (aged 27-76 y; mean, 51.6 ± 13.9 y) with metastatic NENs underwent peptide receptor radionuclide therapy (PRRT) with 177Lu-DOTA-LM3 between August 2017 and December 2019. The median administered activity per cycle was 6.1 ± 0.88 GBq (range, 2.8-7.4 GBq). 68Ga-NODAGA-LM3 PET/CT was used for patient selection and follow-up after 177Lu-DOTA-LM3 PRRT. Morphologic and molecular responses were evaluated in accordance with RECIST 1.1 and the criteria of the European Organisation for Research and Treatment of Cancer (EORTC). Treatment-related adverse events were graded according to the National Cancer Institute Common Terminology Criteria for Adverse Events, version 5.0. Dosimetry was performed on 11 patients and compared with the SSTR agonist 177Lu-DOTATOC in 247 patients undergoing PRRT on the same dosimetry protocol. Results: Higher uptake and a longer effective half-life were found for 177Lu-DOTA-LM3 than for the agonist 177Lu-DOTATOC in the whole body and in the kidneys, spleen, and metastases, resulting in higher mean absorbed organ and tumor doses. All patients tolerated therapy without any serious acute adverse effects. Mild nausea without vomiting was observed in 5 (9.8%) patients; no other symptoms were reported. The most severe delayed adverse event was Common Terminology Criteria (CTC)-3 thrombocytopenia in 3 (5.9%) patients. Neither CTC-4 thrombocytopenia nor CTC-3-4 anemia or leukopenia was observed after treatment. No significant decline in renal function was observed, nor was hepatotoxicity. According to RECIST 1.1, disease control could be reached in 40 patients (disease control rate, 85.1%) of the 47 patients monitored after 177Lu-DOTA-LM3 PRRT, with a partial response in 17 (36.2%) and stable disease in 23 (48.9%), whereas 7 patients (14.9%) had progressive disease, and by EORTC criteria, there was complete remission in 2 patients (4.3%), partial remission in 21 (44.7%), stable disease in 18 (38.3%), and progressive disease in 6 (12.8%). Conclusion: The antagonist PRRT with 177Lu-DOTA-LM3 could be administered without severe adverse effects and was well tolerated by most patients, with thrombocytopenia occurring in only a few. No other severe adverse effects were observed; in particular, there was no nephrotoxicity. The SSTR antagonist 177Lu-DOTA-LM3 appears to be promising for PRRT, provides a favorable biodistribution and higher tumor radiation doses than SSTR agonists, and was effective in treating advanced metastatic NENs, especially in patients with low or no SSTR agonist binding, even achieving complete remission in some patients.

Synthesis, preclinical evaluation, and a pilot clinical imaging study of [18F]AlF-NOTA-JR11 for neuroendocrine neoplasms compared with [68Ga]Ga-DOTA-TATE.

A [18F]AlF-labeled somatostatin receptor (SSTR) antagonist was developed for imaging of neuroendocrine neoplasms (NENs), evaluated and compared with [68Ga]Ga-DOTA-TATE. [18F]AlF-NOTA-JR11 was synthesized manually and qualified with high-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS). The cellular uptake, internalization, and saturation binding were performed with HEK293-SSTR2 cells. Biodistribution and micro-PET imaging were carried out with HEK293-SSTR2 tumor-bearing mice. [18F]AlF-NOTA-JR11 PET/MR imaging and [68Ga]Ga-DOTA-TATE PET/CT were performed with ten patients of NEN at 50~60min post-injection (p.i.). Normal organ biodistribution and tumor detectability were evaluated. [18F]AlF-NOTA-JR11(24~36GBq/μmol) was prepared within 30min and 51.35 ± 3.30% (n > 10)of radiochemical yield. The radiochemical purity was 98.74 ± 1.24% (n > 10). Two stereoisomers were found and confirmed by LC-MS. The cellular uptake of [18F]AlF-NOTA-JR11 and [68Ga]Ga-DOTA-TATE were 4.50 ± 0.31 and 4.50 ± 0.13 %AD/105 cells at 30min, and the internalization at 37°C of [18F]AlF-NOTA-JR11 (5.47 ± 0.32% at 60min) was significantly lower than [68Ga]Ga-DOTA-TATE (66.89 ± 1.62% at 60min). The affinity of [18F]AlF-NOTA-JR11 (Kd = 11.59 ± 1.31nM) was slightly lower than [68Ga]Ga-DOTA-TATE (Kd = 7.36 ± 1.02nM); [18F]AlF-NOTA-JR11 showed high uptake in tumor (9.02 ± 0.92 %ID/g at 60min p.i.) which can be blocked by 50μg of NOTA-JR11 (3.40 ± 1.64 %ID/g at 60min p.i.); the result was coincident with micro-PET imaging. Imaging study of NEN patients showed that more lesions were found only by [18F]AlF-NOTA-JR11 (n = 67 vs. 1 only by [68Ga]Ga-DOTA-TATE), and the uptakes of [18F]AlF-NOTA-JR11 in majority normal organs were significantly lower than [68Ga]Ga-DOTA-TATE. The target to nontarget of maximum of standard uptake value (SUVmax) of [18F]AlF-NOTA-JR11 in liver lesions were significantly higher than those of [68Ga]Ga-DOTA-TATE. Qualitied [18F]AlF-NOTA-JR11 is prepared conveniently with reasonable yield, and it can bind SSTR2 specifically with high affinity. Excellent imaging capability of [18F]AlF-NOTA-JR11 for NENs is superior to [68Ga]Ga-DOTA-TATE, especially in digestive system. It has a great potential for imaging of NENs.

Direct labeling of a somatostatin receptor antagonist via peptide cyclization with Re, 99mTc and 186Re metal centers: Radiochemistry and in vitro evaluation

IntroductionWith the long-term goal of developing a diagnostic (99mTc) and therapeutic (186Re) agent pair for targeting somatostatin receptor (SSTR)-positive neuroendocrine tumors (NETs), we developed novel metal-cyclized peptides through direct labeling of the potent SSTR2 antagonist Ac-4-NO2-Phe-c(DCys-Tyr-DTrp-Lys-Thr-Cys)-DTyr-NH2 (1) with Re (in Re-1), 99mTc (in [99mTc]Tc-1) and 186Re (in [186Re]Re-1). MethodsRe-1 was characterized by LC-ESI-MS and HR-ESI-MS and was tested for receptor affinity in SSTR-expressing cells (AR42J). Radiolabeling of the peptide was achieved via ligand exchange from 99mTc-labeled glucoheptonate or [186Re]ReOCl3(PPh3)2, yielding [99mTc]Tc-1 or [186Re]Re-1, respectively. In vitro stability of [99mTc]Tc-1/[186Re]Re-1 in PBS (10 mM) at pH 7.4 and 37 °C was determined by HPLC analysis. Moreover, [99mTc]Tc-1 stability was tested in cysteine (1 mM) and rat serum under the same conditions. ResultsRe-1 consisted of two isomers, confirmed by LC-ESI-MS, with good SSTR2 affinity (IC50 = 43 ± 6 nM). Optimization of the 99mTc labeling through varying reaction parameters such as pH, reaction time, and Sn2+ and ligand concentrations resulted in high radiochemical yield (RCY ≥92%). Similarly, [186Re]Re-1 was prepared in reasonable RCY (≥50%). Both 99mTc/186Re-tracers consisted of two product isomers as identified by HPLC co-injection with Re-1. While [99mTc]Tc-1 was sufficiently stable in vitro (≥71% intact through 4 h in PBS, cysteine and rat serum), [186Re]Re-1 exhibited more moderate in vitro stability (58% intact after 1 h in PBS). ConclusionsNovel 99mTc/186Re-cyclized SSTR2 antagonist peptides were synthesized and characterized using the Re-cyclized analogue as a reference. Due to the nanomolar SSTR2 affinity of Re-1 and good in vitro stability of [99mTc]Tc-1, the latter shows early promise for development as a radiodiagnostic agent for SSTR-expressing NETs. Advances in knowledge and implications for patient careThe 99mTc-cyclized complex showed promising in vitro properties, and future in vivo studies will determine the potential for translating such a design into the human clinic.

Multimodal Imaging of 2-Cycle PRRT with 177Lu-DOTA-JR11 and 177Lu-DOTATOC in an Orthotopic Neuroendocrine Xenograft Tumor Mouse Model.

Peptide receptor radionuclide therapy (PRRT) using radiolabeled somatostatin receptor (SSTR) analogs is a common approach in advanced neuroendocrine neoplasms. Recently, SSTR antagonists have shown promising results for imaging and therapy due to a higher number of binding sites than in commonly used agonists. We evaluated PRRT with SSTR agonist 177Lu-DOTATOC and antagonist 177Lu-DOTA-JR11 longitudinally in an orthotopic murine pancreatic neuroendocrine neoplasm model expressing human SSTR2. Morphologic and metabolic changes during treatment were assessed using multimodal imaging, including hybrid PET/MRI and SPECT/CT. Methods: In vitro radioligand binding and internalization assays and cell-cycle analysis were performed. SSTR2-transfected BON cells (BON-SSTR2) were used for in vivo experiments. Tumor-bearing mice received 2 intravenous injections of 100 μL of saline, 30 MBq of 177Lu-DOTATOC, or 20 MBq of 177Lu-DOTA-JR11 with an interval of 3 wk. Weekly T2-weighted MRI was performed for tumor monitoring. Viability of the tumor tissue was assessed by 18F-FDG PET/MRI once after PRRT. Tumor and kidney uptake of the respective radiopharmaceuticals was measured 24 h after injection by SPECT/CT. Results: Compared with 177Lu-DOTATOC, 177Lu-DOTA-JR11 treatment resulted in an increased accumulation of cells in G2/M phase. Animals treated with the SSTR antagonist showed a significant reduction in tumor size (P < 0.001) and an increased median survival (207 d; interquartile range [IQR], 132-228) compared with 177Lu-DOTATOC (126 d; IQR, 118-129). SPECT/CT revealed a 4-fold higher median tumor uptake for the antagonist and a 3-fold higher tumor-to-kidney ratio in the first treatment cycle. During the second therapy cycle, tumor uptake of 177Lu-DOTATOC was significantly lower (P = 0.01) whereas 177Lu-DOTA-JR11 uptake remained stable. Imaging of tumor morphology indicated comparatively larger necrotic fractions for 177Lu-DOTA-JR11 despite further tumor growth. These results were confirmed by 18F-FDG PET, revealing the least amount of viable tumor tissue in 177Lu-DOTA-JR11-treated animals, at 6.2% (IQR, 2%-23%). Conclusion:177Lu-DOTA-JR11 showed a higher tumor-to-kidney ratio and a more pronounced cytotoxic effect than did 177Lu-DOTATOC. Additionally, tumor uptake was more stable over the course of 2 treatment cycles.

Somatostatin secretion by Na+-dependent Ca2+-induced Ca2+ release in pancreatic delta-cells.

Pancreatic islets are complex micro-organs consisting of at least three different cell types: glucagon-secreting α-, insulin-producing β- and somatostatin-releasing δ-cells1. Somatostatin is a powerful paracrine inhibitor of insulin and glucagon secretion2. In diabetes, increased somatostatinergic signalling leads to defective counter-regulatory glucagon secretion3. This increases the risk of severe hypoglycaemia, a dangerous complication of insulin therapy4. The regulation of somatostatin secretion involves both intrinsic and paracrine mechanisms5 but their relative contributions and whether they interact remains unclear. Here we show that dapagliflozin-sensitive glucose- and insulin-dependent sodium uptake stimulates somatostatin secretion by elevating the cytoplasmic Na+ concentration ([Na+]i) and promoting intracellular Ca2+-induced Ca2+ release (CICR). This mechanism also becomes activated when [Na+]i is elevated following the inhibition of the plasmalemmal Na+-K+ pump by reductions of the extracellular K+ concentration emulating those produced by exogenous insulin in vivo6. Islets from some donors with type-2 diabetes hypersecrete somatostatin, leading to suppression of glucagon secretion that can be alleviated by a somatostatin receptor antagonist. Our data highlight the role of Na+ as an intracellular second messenger, illustrate the significance of the intraislet paracrine network and provide a mechanistic framework for pharmacological correction of the hormone secretion defects associated with diabetes that selectively target the δ-cells.

Preclinical study of a new 177Lu-labeled somatostatin receptor antagonist in HT-29 human colorectal cancer cells.

Somatostatin receptor-positive neuroendocrine tumors have been targeted using various peptide analogs radiolabeled with therapeutic radionuclides for years. The better biomedical properties of radioantagonists as higher tumor uptake make these radioligands more attractive than agonists for somatostatin receptor-targeted radionuclide therapy. In this study, we tried to evaluate the efficiency of Luthetium-177 (177Lu) radiolabeled DOTA-Peptide 2 (177Lu-DOTA-Peptide 2) as a new radioantagonist in HT-29 human colorectal cancer in vitro and in vivo. DOTA conjugated antagonistic peptide with the sequence of p-Cl-Phe-Cyclo(D-Cys-L-BzThi-D-Aph-Lys-Thr-Cys)-D-Tyr-NH2 (DOTA-Peptide 2) was labeled with 177Lu. In vitro assays (saturation binding assay and internalization test) and animal biodistribution were performed in human colon adenocarcinoma cells (HT-29) and HT-29 tumor-bearing nude mice. 177Lu-DOTA-Peptide 2 showed high stability in acetate buffer and human plasma (>97%). Antagonistic property of 177Lu-DOTA-Peptide 2 was confirmed by low internalization in HT-29 cells (<5%). The desired dissociation constant (Kd =11.14 nM) and effective tumor uptake (10.89 percentage of injected dose per gram of tumor) showed high binding affinity of 177Lu-DOTA-Peptide 2 to somatostatin receptors. 177Lu-DOTA-Peptide 2 demonstrated selective and high binding affinity to somatostatin receptors overexpressed on the surface of HT-29 cancer cells, which could make this radiopeptide suitable for somatostatin receptor-targeted radionuclide therapy.

Design, preparation and biological evaluation of a 177Lu-labeled somatostatin receptor antagonist for targeted therapy of neuroendocrine tumors.

Somatostatin receptor-targeted radionuclide therapy has become an effective treatment in patients with neuroendocrine tumors. Recently, investigations on the development of antagonistic peptides are increasing with possible superior biological properties as opposed to the agonists. Herein, we have reported the development of a new somatostatin receptor peptide ligand labeled with 177Lu to achieve a therapeutic ligand for tumor treatment. The interactions of selected and drown ligands using Avogadro software were docked on somatostatin receptor by Dink algorithm. The best docked peptide-chelator conjugate (DOTA-p-Cl-Phe-Cyclo(d-Cys-l-BzThi-d-Aph-Lys-Thr-Cys)-d-Tyr-NH2) (DOTA-Peptide 2) was synthesized using the Fmoc solid-phase method. DOTA-Peptide 2 was radiolabeled with the 177Lu Trichloride (177LuCl3) solution at 95 °C for 30 min and radiochemical purity (RCP) of 177Lu-DOTA-Peptide 2 solution was monitored by radio-HPLC and radio-TLC procedures. The new radiolabeled peptide was evaluated for stability, receptor binding, internalization, biodistribution and single-photon emission computed tomography (SPECT) imaging using C6 glioma cells and C6 tumor-bearing rats. DOTA-Peptide 2 was obtained with 98% purity and efficiently labeled with 177Lu (RCP > 99%). 177Lu-DOTA-Peptide 2 showed a high value of stability in acetate buffer (91.4% at 312 h) and human plasma (>97% at 24 h). Radioconjugate exhibited low internalization (<5%) and high affinity for somatostatin receptors (Kd = 12.06 nM, Bmax = 0.20 pmol/106 cells) using saturation binding assay. Effective tumor uptake of 7.3% ID/g (percentage of injected dose per gram of tumor) at 4 h post-injection and fast clearance of radiopeptide from blood and other organs led to a high tumor-to-normal organ ratios. SPECT/CT imaging clearly showed the activity localization in tumor. The favorable antagonistic properties of 177Lu-DOTA-Peptide 2 on the somatostatin receptors can make it a suitable candidate for peptide receptor radionuclide therapy (PRRT). In the future study, the therapeutic application of this radiopeptide will be evaluated.

Preparation and microPET imaging of somatostatin receptor antagonist 68Ga-NOTA-JR11

Objective To prepare a novel somatostatin receptor (SSTR) antagonist 68Ga-1, 4, 7-triazacyclononane-1, 4, 7-triacetic acid (NOTA)-JR11 (Cpa-c(D-Cys-Aph(Hor)-D-Aph(Cbm)-Lys-Thr-Cys)-D-Tyr-NH2) tracer and observe its biodistribution and microPET imaging in mice. Methods One ml HCl (0.05 mol/L) containing 68GaCl3 (148 MBq) was added into 65 μl NaAc (1 mol/L) and 4 μg NOTA-JR11. The mixture reacted at 95 ℃ for 15 min, and then was purified with Sep-Pak® C18 Light column to obtain 68Ga-NOTA-JR11. 68Ga-NOTA-JR11 was subjected to quality control analysis including radiochemical purity and in vitro stability by radio-high performance liquid chromatography. Biodistribution of 68Ga-NOTA-JR11 (0.37 MBq) in BALB/c mice (n=9) at 5, 30, 60 min postinjection were observed (n=3 for each time point), and the percentage activity of injection dose per gram of tissue (%ID/g) was calculated. 68Ga-NOTA-JR11 (14.8 MBq) microPET imaging of BALB/c mice (n=1) at 60 min postinjection was observed. Results 68Ga-NOTA-JR11 was obtained successfully within 15 min, with yielding rate of 90%, radiochemical purity of more than 99%, and specific activity of 6.10 GBq/μmol. The tracer showed excellent stability (radiochemical purity: 95%) in different buffers within 150 min. The biodistribution was basically consistent with microPET imaging results. 68Ga-NOTA-JR11 was metabolized through the kidneys and had low uptake in the liver ((0.75±0.26) %ID/g) at 60 min postinjection. Conclusions 68Ga-NOTA-JR11 can be prepared rapidly, with high yielding rate and radiochemical purity. Biodistribution and imaging results provide basic information for the further study of somatostatin receptor imaging in neuroendocrine tumors. Key words: Antistatic agents; Receptors, somatostatin; Isotope labeling; Gallium radioisotopes; Positron-emission tomography; Mice

Treatment of female rhesus macaques with a somatostatin receptor antagonist that increases oocyte fertilization rates without affecting post-fertilization development outcomes.

To determine the effects of PGL1001, a somatostatin receptor isoform-2 (SSTR-2) antagonist, on ovarian follicle development, oocyte fertilization, and subsequent embryo developmental potential in the rhesus macaque. Cycling female rhesus macaques (N= 8) received vehicle through one menstrual (control) cycle, followed by daily injections of PGL1001, a SSTR-2 antagonist, for three menstrual (treatment) cycles. Main endpoints include overall animal health and ovarian hormones (e.g., estradiol [E2], progesterone [P4], and anti-Müllerian hormone [AMH]), ovarian circumference, numbers of oocytes and their maturation status following controlled ovarian stimulation (COS), as well as oocyte fertilization and subsequent blastocyst rates that were assessed in control and PGL1001 treatment cycles. Circulating PGL1001 levels were assessed at baseline as well as 6, 60, and 90days during treatment. PGL1001 treatment did not impact overall animal health, menstrual cycle length, or circulating levels of ovarian hormones (E2, P4, and AMH) in comparison to vehicle treatment during natural cycles. PGL1001 treatment increased (p˂0.05) ovarian circumference and the day 8 to day 1 ratio of AMH levels (p˂0.05) during a COS protocol, as well as oocyte fertilization rates compared to the vehicle treatment interval. Blastocyst development rates were not significantly different between vehicle and PGL1001 treatment groups. Prolonged treatment with PGL1001 appears to be safe and does not affect rhesus macaque general health, menstrual cycle length, or ovarian hormone production. Interestingly, PGL1001 treatment increased the fertilization rate of rhesus macaque oocytes collected following ovarian stimulation.

NOTA and NODAGA [99mTc]Tc- and [186Re]Re-Tricarbonyl Complexes: Radiochemistry and First Example of a [99mTc]Tc-NODAGA Somatostatin Receptor-Targeting Bioconjugate.

With the long-term goal of developing theranostic agents for applications in nuclear medicine, in this work we evaluated the well-known NOTA and NODAGA chelators as bifunctional chelators (BFCs) for the [99mTc/186Re]Tc/Re-tricarbonyl core. In particular, we report model complexes of the general formula fac-[M(L)(CO)3]+ (M = Re, 99mTc, 186Re) where L denotes NOTA-Pyr (1) or NODAGA-Pyr (2), which are derived from conjugation of NOTA/NODAGA with pyrrolidine (Pyr). Further, as proof-of-principle, we synthesized the peptide bioconjugate NODAGA-sst2-ANT (3) and explored its complexation with the fac-[Re(CO)3]+ and fac-[99mTc][Tc(CO)3]+ cores; sst2-ANT denotes the somatostatin receptor (SSTR) antagonist 4-NO2-Phe-c(DCys-Tyr-DTrp-Lys-Thr-Cys)-DTyr-NH2. Rhenium complexes Re-1 through Re-3 were synthesized and characterized spectroscopically, and receptor binding affinity was demonstrated for Re-3 in SSTR-expressing cells (AR42J, IC50 = 91 nM). Radiolabeled complexes [99mTc]Tc/[186Re]Re-1/2 and [99mTc]Tc-3 were prepared in high radiochemical yield (>90%, determined by radio-HPLC) by reacting [99mTc]/[186Re][Tc/Re(OH2)3(CO)3]+ with 1-3 and correlated well with the respective Re-1 through Re-3 standards in comparative HPLC studies. All radiotracers remained intact through 24 h (99mTc-labeled complexes) or 48 h (186Re-labeled complexes) against 1 mM l-histidine and 1 mM l-cysteine (pH 7.4, 37 °C). Similarly, rat serum stability studies displayed no decomposition and low nonspecific binding of 9-24% through 4 h. Biodistribution of [99mTc]Tc-3 in healthy CF-1 mice demonstrated a favorable pharmacokinetic profile. Rapid clearance was observed within 1 h post-injection, predominantly via the renal system (82% of the injected dose was excreted in urine by 1 h), with low kidney retention (% ID/g: 11 at 1 h, 5 at 4 h, and 1 at 24 h) and low nonspecific uptake in other organs/tissues. Our findings establish NOTA and NODAGA as outstanding BFCs for the fac-[M(CO)3]+ core in the design and development of organometallic radiopharmaceuticals. Future in vivo studies of [99mTc]Tc- and [186Re]Re-tricarbonyl complexes of NODAGA/NOTA-biomolecule conjugates will further probe the potential of these chelates for nuclear medicine applications in diagnostic imaging and targeted radiotherapy, respectively.

Cortistatin regulates glucose-induced electrical activity and insulin secretion in mouse pancreatic beta-cells.

Although there is growing evidence that cortistatin regulates several functions in different tissues, its role in the endocrine pancreas is not totally known. Here, we aim to study the effect of cortistatin on pancreatic beta-cells and glucose-stimulated insulin secretion (GSIS). Exposure of isolated mouse islets to cortistatin inhibited GSIS. This effect was prevented using a somatostatin receptor antagonist. Additionally, cortistatin hyperpolarized the membrane potential and reduced glucose-induced action potentials in isolated pancreatic beta-cells. Cortistatin did not modify ATP-dependent K+ (KATP) channel activity. In contrast, cortistatin increased the activity of a small conductance channel with characteristics of G protein-coupled inwardly rectifying K+ (GIRK) channels. The cortistatin effects on membrane potential and GSIS were largely reduced in the presence of a GIRK channel antagonist and by down-regulation of GIRK2 with small interfering RNA. Thus, cortistatin acts as an inhibitory signal for glucose-induced electrical activity and insulin secretion in the mouse pancreatic beta-cell.

SSTR-Mediated Imaging in Breast Cancer: Is There a Role for Radiolabeled Somatostatin Receptor Antagonists?

Recent studies have shown enhanced tumor targeting by novel somatostatin receptor (SSTR) antagonists compared with clinically widely used agonists. However, these results have been obtained mostly in neuroendocrine tumors, and only limited data are available for cancer types with lower SSTR expression, including breast cancer (BC). To date, two studies have reported higher binding of the antagonist than the agonist in BC, but in both studies only a limited number of cases were evaluated. In this preclinical study, we further investigated whether the application of an SSTR antagonist can improve SSTR-mediated BC imaging in a large panel of BC specimens. We also generated an in vivo BC mouse model and performed SPECT/MRI and biodistribution studies. Methods: Binding of 111In-DOTA-Tyr3-octreotate (SSTR agonist) and 111In-DOTA-JR11 (SSTR antagonist) to 40 human BC specimens was compared using in vitro autoradiography. SSTR2 immunostaining was performed to confirm SSTR2 expression of the tumor cells. Furthermore, binding of the radiolabeled SSTR agonist and antagonist was analyzed in tissue material from 6 patient-derived xenografts. One patient-derived xenograft, the estrogen receptor-positive model T126, was chosen to generate in vivo mouse models containing orthotopic breast tumors for in vivo SPECT/MRI and biodistribution studies after injection with 177Lu-DOTA-Tyr3-octreotate or 177Lu-DOTA-JR11. Results:111In-DOTA-JR11 binding to human BC tissue was significantly higher than 111In-DOTA-Tyr3-octreotate binding (P < 0.001). The median ratio of antagonist binding versus agonist binding was 3.39 (interquartile range, 2-5). SSTR2 immunostaining confirmed SSTR2 expression on the tumor cells. SPECT/MRI of the mouse model found better tumor visualization with the antagonist. This result was in line with the significantly higher tumor uptake of the radiolabeled antagonist than of the agonist as measured in biodistribution studies 285 min after radiotracer injection (percentage injected dose per gram of tissue: 1.92 ± 0.43 vs. 0.90 ± 0.17; P = 0.002). Conclusion: SSTR antagonists are promising candidates for BC imaging.

Synthesis and evaluation of a 99mTc tricarbonyl-labeled somatostatin receptor-targeting antagonist peptide for imaging of neuroendocrine tumors

IntroductionA somatostatin receptor (SSTR)-targeting antagonist peptide (sst2-ANT) was radiolabeled with 99mTc tricarbonyl via a tridentate [N,S,N]-type ligand (L) to develop a radiodiagnostic agent, 99mTcL-sst2-ANT, for imaging of SSTR-expressing neuroendocrine tumors. MethodsReceptor affinity was assessed in vitro with the nonradioactive analogue, ReL-sst2-ANT, via a challenge experiment in AR42J cells with 125I–SS-14 as the competing radioligand. Preparation of 99mTcL-sst2-ANT was achieved via reaction of [99mTc(CO)3(H2O)3]+ with L-sst2-ANT. To test the stability of the radiolabeled complex, challenge experiments were performed in phosphate-buffered saline solutions containing cysteine or histidine and also in mouse serum. Biodistribution and micro-SPECT/CT imaging studies were performed in AR42J tumor-bearing female ICR SCID mice. ResultsThe half maximal inhibitory concentration (IC50 value) of ReL-sst2-ANT in AR42J cells was 15nM. Preparation of 99mTcL-sst2-ANT was achieved with ≥97% radiochemical yield (RCY) and was verified by HPLC co-elution with the ReL-sst2-ANT analogue. The radiolabeled complex remained intact for up to 24h in high concentration solutions of cysteine and histidine at 37°C. Furthermore, the radiotracer was 90% stable for 1h at 37°C in mouse serum. Micro-SPECT/CT images showed clear uptake in tumors and were supported by the biodistribution data, in which the 3.2% ID/g tumor uptake at 4h was significantly blocked by co-administration of nonradioactive SS-14. ConclusionsA [99mTc(CO)3(N,S,N)]+ chelate was employed for radiolabeling of an SSTR-targeting antagonist peptide. Synthesis of 99mTcL-sst2-ANT was achieved in high RCY, and the resulting complex displayed high in vitro stability. Somatostatin receptor affinity was retained in both cells and in tumor-bearing mice, where the complex successfully targeted SSTR-positive tumors via a receptor-mediated process.Advances in Knowledge and Implications for Patient Care.This first 99mTc-tricarbonyl-labeled SSTR antagonist peptide showed promising in vivo tumor targeting in mice. Future studies may lead to translation of a similar design into the clinic.